Morphologic features of large choroidal vessel layer: age-related macular degeneration, polypoidal choroidal vasculopathy, and central serous chorioretinopathy.

PURPOSE: To compare choroidal vascular characteristics of age-related macular degeneration (AMD), polypoidal choroidal vasculopathy (PCV), and central serous chorioretinopathy (CSC) by qualitative and quantitative analyses using swept-source en face optical coherence tomographic (OCT) images. METHODS: Eyes with non-neovascular AMD (n = 32), neovascular AMD (n = 30), thick and thin choroid PCV (n = 33 and 27), and CSC (n = 34) were enrolled. Subfoveal choroidal thickness (SFCT) and the presence and patterns of pachyvessels were assessed. En face images of the large choroidal vessel layer were converted to binary images for the analysis of vascular density. RESULTS: Pachyvessels were identified in 8 (25%), 14 (46%), 28 (85%), 26 (96%), and 34 (100%) non-neovascular AMD, neovascular AMD, thin choroid PCV, thick choroid PCV, and CSC eyes, respectively (P < 0.001). The pattern of pachyvessels was focal in non-neovascular AMD (100%), neovascular AMD (79%), and thin choroid PCV (89%) while the pattern was mostly diffuse in CSC (88%) and thick choroid PCV (81%). The mean choroidal vascular density in a 6 × 6 mm2 macular area of each group was 45.3%, 46.9%, 47.0%, 52.5%, and 54.8%, respectively (P < 0.001). Post hoc analysis revealed significantly higher vascular density in CSC compared with other types (all P < 0.001) except PCV with thick choroid (P = 0.066). CONCLUSIONS: Similarities in vascular density of the large choroidal vessel layer and pachyvessel pattern were between CSC and thick choroid PCV and between AMD and thin choroid PCV, suggesting common pathophysiology involving choroidal changes in these eyes.

Enhancing proliferation and optimizing the culture condition for human bone marrow stromal cells using hypoxia and fibroblast growth factor-2.

This study aimed to determine the cellular characteristics and behaviors of human bone marrow stromal cells (hBMSCs) expanded in media in a hypoxic or normoxic condition and with or without fibroblast growth factor-2 (FGF-2) treatment. hBMSCs isolated from the vertebral body and expanded in these four groups were evaluated for cellular proliferation/migration, colony-forming units, cell-surface characterization, in vitro differentiation, in vivo transplantation, and gene expression. Culturing hBMSCs using a particular environmental factor (hypoxia) and with the addition of FGF-2 increased the cellular proliferation rate while enhancing the regenerative potential, modulated the multipotency-related processes (enhanced chondrogenesis-related processes/osteogenesis, but reduced adipogenesis), and increased cellular migration and collagen formation. The gene expression levels in the experimental samples showed activation of the hypoxia-inducible factor-1 pathway and glycolysis in the hypoxic condition, with this not being affected by the addition of FGF-2. The concurrent application of hypoxia and FGF-2 could provide a favorable condition for culturing hBMSCs to be used in clinical applications associated with bone tissue engineering, due to the enhancement of cellular proliferation and regenerative potential.

Novel Technique for Isolating Human Bone Marrow Stem Cells Using Hyaluronic Acid Hydrogel.

Centrifugation based on density gradients is a general methodology for isolating human bone marrow (hBM)-derived mesenchymal stem cells (hBMSCs). The mononuclear cell (MNC) layer can be obtained using a density gradient solution in the conventional protocol, but it is not suitable for direct transplantation due to the possible toxicity of this solution. The results obtained are also influenced by the skill level when applying the technique, which involves time-consuming processes. We have developed a novel protocol for isolating hBMSCs using hyaluronic acid (HA), which is the most widely used injectable biomaterial in clinical settings and a major component of the extracellular matrix. Laying hBM over the HA and then applying centrifugation yielded three separate layers, with the HA layer, including MNCs being the most superficial one. Increasing the volume of HA and/or its crosslinking rate enhanced the yield of MNCs from hBM, and the cell yield was also significantly higher for a lower centrifugal acceleration (530 g) than for a higher one (1500 g). Isolated hBMSCs by HA exhibited similar biological characteristics such as in terms of their proliferation rate, fibroblast-like morphology, cell-cycle status, immunophenotype, and multipotency. The use of either type of hBMSC confirmed the regenerative potential of bone and bone marrow-like tissue in ectopic transplantation models. This is the first report of a novel protocol for isolating hBMSCs that utilize HA. We suggest that this novel isolation technique can be used for the direct application of autogenous MSCs with advantages of being less time-consuming and involving steps that are easier to perform.

Autogenous Mesenchymal Stem Cells from the Vertebral Body Enhance Intervertebral Disc Regeneration via Paracrine Interaction: An in Vitro Pilot Study.

Several in vivo studies have found that transplanting mesenchymal stem cells (MSCs) into degenerative intervertebral discs (IVDs) leads to regeneration of disc cells. Since the exact underlying mechanisms are not understood, we investigated the mechanisms of action of MSCs in regeneration of degenerative IVDs via paracrine actions. Human MSCs and degenerative disc cells from the same donor vertebrae were directly or indirectly cocultured. The multidifferentiation potential, cell proliferation, collagen synthesis, and mRNA expression levels were assessed. The proliferation rates of MSCs and degenerative disc cells were higher in the coculture system than in the monolayer cultures or in the conditioned medium of each cell type. During coculturing with nucleus pulposus (NP) cells, mRNA expression of the extracellular matrix (ECM) components aggrecan, versican (VCAN), SOX9, and type II and type VI collagen was significantly increased in MSCs, whereas mRNA expression for type V collagen was increased in MSCs cocultured with annulus fibrosus (AF) cells. In addition, the accumulation of total ECM collagen was greater in cocultured degenerative disc cells than in monocultured cells. During coculturing, MSCs downregulated the expression levels of various proinflammatory cytokine genes in degenerative NP [interleukin-1α ( IL-1α), IL-1ß, IL-6, and tumor necrosis factor-α ( TNF-α)] and AF cells ( IL-1α and IL-6), which are involved in the degradation of ECM molecules. In association with the trophic effect of MSCs on degenerative disc cells, upregulation of growth factor mRNA expression was shown in MSCs cocultured with degenerative NP cells [epidermal growth factor ( EGF), insulin-like growth factor-1 ( IGF-1), osteogenic protein-1 ( OP-1), growth and differentiation factor-7 ( GDF-7), and transforming growth factor-ß ( TGF-ß)] or degenerative AF cells ( IGF-1, OP-1, and GDF-7). In terms of MSC-based clinical approaches to IVD regeneration, implanting MSCs into a degenerative IVD may both stimulate MSC differentiation into an NP- or AF-like phenotype and stimulate the biological activation of degenerative disc cells for self-repair.

Human bone marrow stem cells cultured under hypoxic conditions present altered characteristics and enhanced in vivo tissue regeneration.

Human bone marrow mesenchymal stem cells (hBMSCs) were isolated from bone marrow of the vertebral body. The hBMSCs were cultured under either hypoxic (1% O2) or normoxic (21% O2; control) conditions and the characteristics as mesenchymal stem cells were compared. Results revealed that hypoxia reduced proliferative potential and colony-forming efficiency of hBMSCs, and significantly enhanced osteogenic and chondrogenic differentiation. The hBMSCs enhanced the regenerative potential of bone in vivo. In vitro synthesis of soluble and insoluble collagen was significantly increased in the hypoxic condition. In vivo collagen tissue regeneration was also enhanced under the hypoxic condition, with concomitant increased expressions of various subtypes of collagen and lysyl-oxidase family mRNA. MicroRNA assays revealed that miR-155-5p, which negatively regulates HIF-1α, was significantly highly expressed. These observations demonstrate that hBMSCs obtained from human vertebrae exhibit altered characteristics under hypoxic conditions, and each factor contributing to hBMSC-mediated tissue healing should be evaluated with the goal of allowing their clinical application.

Effect of FGF-2 on collagen tissue regeneration by human vertebral bone marrow stem cells.

The effects of fibroblast growth factor-2 (FGF-2) on collagen tissue regeneration by human bone marrow stem cells (hBMSCs) were investigated. hBMSCs were isolated from human vertebral body bone marrow during vertebral surgery and a population of hBMSCs with the characteristics of mesenchymal stem cells was observed. The FGF-2 treatment (5 ng/mL) affected on the colony-forming efficiency, proliferation, and in vitro differentiation of hBMSCs. Insoluble/soluble collagen and hydroxyproline synthesis was significantly enhanced in hBMSCs expanded with FGF-2 and the treatment of FGF-2 caused a reduction in the mRNA expression of collagen type I, but an increase of collagen types II and III along with lysyl oxidase family genes. Collagen formation was also examined using an in vivo assay model by transplanting hBMSCs into immunocompromised mice (n=4) and the histologic and immunohistochemical results revealed that significantly more collagen with a well-organized structure was formed by FGF-2-treated hBMSCs at 8 weeks posttransplantation (P<0.05). The DNA microarray assay demonstrated that genes related to extracellular matrix formation were significantly upregulated. To elucidate the underlying mechanism, chemical inhibitors against extracellular-signal-regulated kinase (ERK) and phosphoinositide 3-kinase (PI3K) were treated and following downstream expression was observed. Collectively, FGF-2 facilitated the collagen-producing potency of hBMSCs both in vitro and in vivo, rendering them more suitable for use in collagen regeneration in the clinical field.

Differential effect of water-soluble chitin on collagen synthesis of human bone marrow stem cells and human periodontal ligament stem cells.

Human bone marrow stem cells (hBMSCs) represent a promising regenerative material because of their mutipotency, including their ability to regenerate collagenous soft tissues. We previously found that water-soluble chitin (WSC) enhances the ability of human periodontal ligament stem cells (hPDLSCs) to synthesize collagen tissue. The aim of this study was to determine the effects of WSC on hBMSCs and hPDLSCs for the collagen synthesis both in vitro and in vivo. hBMSCs and hPDLSCs were isolated and expanded with or without 0.3 mg/mL WSC. A series of in vitro and in vivo analyses were performed to evaluate their characteristics as stem cell populations. Then, collagen and hydroxyproline assays were conducted using both in vitro and in vivo assay models, and the real-time polymerase chain reaction was performed to analyze the expression of collagen-related markers. WSC-treated and nontreated hBMSCs and hPDLSCs were transplanted into immunocompromised mice, and histology and immunohistochemistry analyses were conducted after 8 weeks. The in vitro results showed that those cells possessed the characteristics of mesenchymal stem cells. The amount of soluble collagen synthesized was significantly greater in WSC-treated hBMSCs than in the nontreated group; conversely, treatment of hPDLSCs with WSC decreased the formation of soluble collagen. The amount of insoluble collagen synthesized was greater in the WSC-treated groups than in the nontreated groups for both hBMSCs and hPDLSCs. The hydroxyproline contents of the regenerated soluble and insoluble collagens were similar. The expressions of mRNA for collagen types I-V, hyaluronic acid synthase 1 (HAS1), HAS2, and HAS3, and the LOX family were higher in WSC-treated hPDLSCs than in the nontreated group, whereas WSC increased the expression of collagen type III and decreased that of collagen type I in hBMSCs. The histology and immunohistochemistry results revealed that WSC significantly increased the amount of collagen formed in vivo by both types of stem cells. Collectively, treatment with WSC significantly enhanced the collagen-forming potentials of hBMSCs and hPDLSCs, but the collagen they produced exhibited distinctively different characteristics. These findings suggest that the appropriate stem-cell source should be chosen based on the purpose of the required regenerated tissue.

Spinal cord herniation after multilevel anterior cervical corpectomy and fusion for ossification of the posterior longitudinal ligament of the cervical spine.

The authors report the case of a 52-year-old man who had undergone resection of an ossified posterior longitudinal ligament via the anterior approach. The patient experienced postoperative neurological deterioration that may have been caused by a massive cord herniation associated with a dural defect at the corpectomy site. Spinal cord herniation may develop as a complication of anterior cervical decompression. Surgeons should be alert to this condition when planning to treat cervical ossification of the ossified posterior longitudinal ligament via the anterior approach.

The clinical characteristics and risk factors for the adjacent segment degeneration in instrumented lumbar fusion.

STUDY DESIGN: A retrospective study. OBJECTIVE: The aims of this study were to evaluate the clinical significance of, characteristics of, and risk factors for adjacent segment degeneration (ASD) in patients who have undergone instrumented lumbar fusion. SUMMARY OF BACKGROUND DATA: ASD has been considered a potential long-term complication of spinal arthrodesis. However, the exact mechanisms and risk factors related to ASD are not completely understood. METHODS: A total of 48 patients who underwent instrumented lumbar fusion at L4-5 and had minimal ASD preoperatively were evaluated. The patients were divided into 2 groups at follow-up according to the development of ASD defined by radiologic criteria. Through review of their medical records and the radiologic files, the following variables were evaluated in the 2 groups: basic demographic data, body weight, body height, body mass index, bone mineral density, types of surgical approaches, preoperative and postoperative segmental and lumbar lordosis, and clinical outcomes. RESULTS: ASD was found in 30 (62.5%) patients. The variables that showed statistical intergroup differences were the mean age at surgery, the mean difference in the degree of preoperative from postoperative lumbar lordosis, and the proportion of patients who underwent anterior lumbar interbody fusion. However, there were no statistically significant intergroup differences in the Japanese Orthopedic Association score at 1-year postoperatively or at the final follow-up, or in the recovery rate, success rate, and complication rate. CONCLUSIONS: Radiographic ASD is relatively common long-term finding associated with instrumented lumbar fusion. However, radiographic evidence of ASD does not necessarily correlate with a poor outcome. Our results suggest that advanced age, anterior lumbar interbody fusion, and the restoration of the preoperative standing lumbar lordosis may have a protective effect against the development of ASD.

Prospective comparison of the 'inside-out' and 'outside-in' transobturator-tape procedures for the treatment of female stress urinary incontinence.

The aim of this study was to prospectively compare the efficacy and safety of 'inside-out' (TVT-O) and 'outside-in' (TOT) transobturator tape procedures for treatment of female stress urinary incontinence (SUI). One hundred women with SUI were alternately assigned to TVT-O (n = 50) or TOT (n = 50). About 1 year after surgery, we compared surgical outcomes in the two groups. TVT-O and TOT showed similar rates of cure (86 vs 92%). Approximately 1 year after surgery, Incontinence Quality of Life questionnaire parameters improved significantly in both groups (p < 0.05), but the two groups did not differ (p > 0.05). The rates of patient satisfaction with TVT-O and TOT (96 vs 98%) were similar. These preliminary results suggest that TVT-O and TOT are equally effective and safe procedures for women with SUI. However, this study was unable to identify a difference between the two procedures due to the underpowered nature of the study.

Outcomes following repeat mid urethral synthetic sling after failure of the initial sling procedure: rediscovery of the tension-free vaginal tape procedure.

PURPOSE: We evaluated outcomes of the repeat mid urethral sling to treat recurrent or persistent stress urinary incontinence after failure of an initial mid urethral sling. MATERIALS AND METHODS: We retrospectively analyzed data on patients who underwent the repeat mid urethral sling procedure due to persistent or recurrent stress urinary incontinence. Repeat slings were placed without removal of the previous sling. All patients were followed at least 1 year after the second mid urethral sling. RESULTS: Of the 31 female patients with a repeat mid urethral sling 29 were followed, including 13 with a retropubic and 16 with a transobturator sling. For the first mid urethral sling 17 patients received a retropubic sling (tension-free vaginal tape) and 12 received a transobturator sling (6 inside out and 6 outside in procedures). Cure and improvement rates irrespective of the approach were 75.9% (22 of 29 patients) and 6.9% (2 of 29), respectively. Cure rates for the retropubic and transobturator slings were 92.3% (12 of 13 patients) and 62.5% (10 of 16), respectively, a difference that did not quite attain statistical significance (p = 0.089). CONCLUSIONS: The repeat mid urethral sling for persistent or recurrent stress urinary incontinence has a lower cure rate than the initial sling. However, the retropubic approach tends to have a higher cure rate than the transobturator approach in repeat sling cases.