Comparison of the Anti-Inflammatory Effects of Mouse Adipose- and Bone-Marrow-Derived Multilineage-Differentiating Stress-Enduring Cells in Acute-Phase Spinal Cord Injury.

Abstract Spinal cord injury (SCI) is a serious neurological disorder, with the consequent disabilities conferred by this disorder typically persisting for life. Multilineage-differentiating stress-enduring (Muse) cells are endogenous stem cells that can be collected from various tissues as well as from mesenchymal stem cells (MSCs); additionally, these Muse cells are currently being used in clinical trials. The anti-inflammatory effect of stem cell transplantation prevents secondary injuries of SCI; however, its effect on Muse cells remains unclear. In this study, we aimed to compare the anti-inflammatory effects of adipose (AD)- and bone marrow (BM)-Muse cells that were isolated from mice (6-week-old C57BL/6J) following intralesional administration during the acute phase of SCI. Flow cytometry was used to isolate Muse cells from AD and BM MSCs. The percentage of Muse cells was 3.9 and 2.7% for AD and BM MSCs, respectively. To examine cell viability, Muse cells were incubated under H2O2-induced oxidative stress conditions. Overall, AD-Muse cells exhibited higher viability than BM-Muse cells (p = 0.032). In enzyme-linked immunosorbent assay analysis, AD-Muse cells displayed greater secretion of brain-derived neurotrophic factor (BDNF; p = 0.008), vascular endothelial growth factor (p = 0.032), and hepatocyte growth factor (p = 0.016). DNA microarray analysis revealed higher expression of Bdnf, neurotrophin-3 (Ntf3), nerve growth factor (Ngf), pleiotrophin (Ptn), and midkine (Mdk) in AD-Muse cells than in BM-Muse cells. To assess their anti-inflammatory effects in vitro, Muse cells and macrophages were co-cultured, and the levels of cytokines (tumor necrosis factor [TNF] α and interleukin [IL] 10) were measured in the medium. Consequently, we found that TNFα levels were lower in AD-Muse cells than in BM-Muse cells (p = 0.009), and IL10 levels were higher in AD-Muse cells than in BM-Muse cells (p = 0.008). Further, we induced moderate injuries via contusion of the spinal cord at the T10 level; Muse cells were transplanted intralesionally 7 days post-SCI. The number of surviving cells, alongside the number of CD86+ (M1 inflammatory effect), and CD206+ (M2 anti-inflammatory effect) macrophages in the spinal cord were measured 7 days post-transplantation. The number of surviving AD-Muse cells was higher than the number of surviving BM-Muse cells (ratio of AD-Muse/BM-Muse = 2.5, p > 0.05). The M1/M2 ratio in the AD-Muse cell-group (0.37) was lower than that in the control (phosphate-buffered saline) group (3.60, p = 0.008). The lesion area in the AD-Muse cell group was smaller than that in the BM-non-Muse (p = 0.049) and control groups (p = 0.012). As AD-Muse cells conferred a higher cell survival and neurotrophic factor secretion ability in vitro, AD-Muse cells demonstrated reduced inflammation after SCI. Overall, intralesional AD-Muse cell therapy is a potential therapeutic candidate that is expected to exhibit anti-inflammatory effects following acute SCI.

Intraperitoneal Administration of Etizolam Improves Locomotor Function in Mice After Spinal Cord Injury.

Neuroinflammation occurs in the acute phase of spinal cord injury (SCI) and inhibits neural regeneration. In mouse models, etizolam (ETZ) is a strong anxiolytic with unclear effects on SCI. This study investigated the effects of short-term administration of ETZ on neuroinflammation and behavior in mice after SCI. We administrated an ETZ (0.5 mg/kg) daily intraperitoneal injection from the day after SCI for 7 days. Mice were randomly divided into three groups (sham group: only laminectomy, saline group, and ETZ group). Inflammatory cytokine concentrations in the injured spinal cord epicenter were measured using an enzyme-linked immunosorbent assay on day 7 after SCI to evaluate spinal cord inflammation in the acute phase. Behavior analysis was performed the day before surgery and on days 7, 14, 28, and 42 after surgery. The behavioral analysis included anxiety-like behavior using the open field test, locomotor function using the Basso Mouse Scale, and sensory function using the mechanical and heat test. Inflammatory cytokine concentrations were significantly lower in the ETZ group than in the saline group in the acute phase after spinal surgery. After SCI, anxiety-like behaviors and sensory functions were comparable between the ETZ and saline groups. ETZ administration reduced neuroinflammation in the spinal cord and improved locomotor function. Gamma-amino butyric acid type A receptor stimulants may be effective therapeutic agents for patients with SCI.

Association between Preoperative Urine Culture and Urinary Tract Infection after Spinal Surgery.

STUDY DESIGN: This is a retrospective study. PURPOSE: This study assessed risk factors accounting for urinary tract infections (UTIs) to determine whether preoperative asymptomatic UTI (aUTI) could be used to predict UTIs in patients after spinal surgery. OVERVIEW OF LITERATURE: UTI is a spinal surgery complication that increases the incidence of surgical site infections. However, the risk factors for UTIs after spinal surgery remain unclear. METHODS: This study included 509 (mean age, 54.5 years; 239 males and 270 females) patients who underwent posterior spine surgery at the department of the current study. First, clean catch urine was collected, after which a urine culture was performed for all patients before surgery. Preoperative detection of the aUTI (>105 colony-forming units/mL) rate was then determined. Subsequently, risk factors for postoperative UTI were evaluated using logistic regression analysis with the following as independent variables: age, sex, obesity, diabetes, spinal cord tumor, the preoperative Japanese Orthopedic Association (JOA) score, JOA-bladder function, preoperative urine culture positivity, aUTI, preoperative Escherichia coli detection, the postoperative catheter placement period, instrumentation, number of surgical levels, surgery duration, and blood loss. RESULTS: The preoperative aUTI and postoperative UTI incidences were 8.1% and 4.1%, respectively. Furthermore, multivariate logistic analysis showed that the risk factor for postoperative UTI was preoperative aUTI (odds ratio, 4.234; 95% confidence interval, 1.532-11.702; p=0.005). CONCLUSIONS: Preoperative aUTI is a risk factor for UTI in patients after spinal surgery.

Protecting Surgeons' Fingers from Radiation Exposure during Lumbosacral Selective Nerve Root Block.

INTRODUCTION: Fluoroscopy-guided selective nerve root block (SNRB) is useful for the diagnosis and treatment of nerve root pain. However, the procedure exposes the surgeon's hands to radiation. Therefore, the purpose of this randomized prospective study was to assess the radiation exposure per unit time of the surgeon's fingers during performance of a lumbosacral SNRB and to calculate the annual exposure time limits for four hand-protection methods. METHODS: We prospectively recruited patients scheduled for an SNRB and measured the radiation exposure using a ring-type passive radiation dosimetry device attached to the distal phalanx of the index finger of the hand performing the needle placement. Patients were randomly divided into the following four groups: a) the direct exposure group, b) the 0.03-mmPb glove group, c) the 0.25-mmPb glove group, and d) the forceps group (in which the needle was held using forceps such that the fingers did not enter the irradiation field). RESULTS: We recruited 40 consecutive patients (16 men and 24 women), with a mean age of 69 years. In all cases, SNRB was successfully performed without complications. The average exposure per hour for each of the four groups was as follows: 0.67 ± 0.56 mSv/s in the direct exposure group, 0.12 ± 0.07 mSv/s in the 0.03-mmPb glove group, 0.019 ± 0.02 mSv/s in the 0.25-mmPb glove group, and 0.001 ± 0.004 mSv/s in the forceps group (p < 0.01). The average annual exposure time limit was 12.4 min in the direct exposure group, 67.9 min in the 0.03-mmPb glove group, 7.5 h in the 0.25-mmPb glove group, and 5.0 days in the forceps group. CONCLUSIONS: Using a radiation reduction glove or forceps greatly decreased the radiation exposure and increased the annual exposure time limit for SNRB.

Neurotrophic Factor Secretion and Neural Differentiation Potential of Multilineage-differentiating Stress-enduring (Muse) Cells Derived from Mouse Adipose Tissue.

Multilineage-differentiating stress-enduring (Muse) cells are endogenous pluripotent stem cells that can be isolated based on stage-specific embryonic antigen-3 (SSEA-3), a pluripotent stem cell-surface marker. However, their capacities for survival, neurotrophic factor secretion, and neuronal and glial differentiation are unclear in rodents. Here we analyzed mouse adipose tissue-derived Muse cells in vitro. We collected mesenchymal stem cells (MSCs) from C57BL/6 J mouse adipose tissue and separated SSEA-3+, namely Muse cells, and SSEA-3-, non-Muse cells, to assess self-renewability; pluripotency marker expression (Nanog, Oct3/4, Sox2, and SSEA-3); spontaneous differentiation into endodermal, mesodermal, and ectodermal lineages; and neural differentiation capabilities under cytokine induction. Neurally differentiated Muse and non-Muse cell functions were assessed by calcium imaging. Antioxidant ability was measured to assess survival under oxidative stress. Brain-derived neurotrophic factor (BDNF), vascular endothelial cell growth factor (VEGF), and hepatocyte growth factor (HGF) secretion were analyzed in enzyme-linked immunosorbent assays. SSEA-3+ Muse cells (6.3 ± 1.9% of mouse adipose-MSCs), but not non-Muse cells, exhibited self-renewability, spontaneous differentiation into the three germ layers, and differentiation into cells positive for Tuj-1 (27 ± 0.9%), O4 (17 ± 3.4%), or GFAP (23 ± 1.3%) under cytokine induction. Neurally differentiated Muse cells responded to KCl depolarization with greater increases in cytoplasmic Ca2+ levels than non-Muse cells. Cell survival under oxidative stress was significantly higher in Muse cells (50 ± 2.7%) versus non-Muse cells (22 ± 2.8%). Muse cells secreted significantly more BDNF, VEGF, and HGF (273 ± 12, 1479 ± 7.5, and 6591 ± 1216 pg/mL, respectively) than non-Muse cells (133 ± 4.0, 1165 ± 20, and 2383 ± 540 pg/mL, respectively). Mouse Muse cells were isolated and characterized for the first time. Muse cells showed greater pluripotency-like characteristics, survival, neurotrophic factor secretion, and neuronal and glial-differentiation capacities than non-Muse cells, indicating that they may have better neural-regeneration potential.