Sodium oligomannate alters gut microbiota, reduces cerebral amyloidosis and reactive microglia in a sex-specific manner.

It has recently become well-established that there is a connection between Alzheimer's disease pathology and gut microbiome dysbiosis. We have previously demonstrated that antibiotic-mediated gut microbiota perturbations lead to attenuation of Aß deposition, phosphorylated tau accumulation, and disease-associated glial cell phenotypes in a sex-dependent manner. In this regard, we were intrigued by the finding that a marine-derived oligosaccharide, GV-971, was reported to alter gut microbiota and reduce Aß amyloidosis in the 5XFAD mouse model that were treated at a point when Aß burden was near plateau levels. Utilizing comparable methodologies, but with distinct technical and temporal features, we now report on the impact of GV-971 on gut microbiota, Aß amyloidosis and microglial phenotypes in the APPPS1-21 model, studies performed at the University of Chicago, and independently in the 5X FAD model, studies performed at Washington University, St. Louis.Methods To comprehensively characterize the effects of GV-971 on the microbiota-microglia-amyloid axis, we conducted two separate investigations at independent institutions. There was no coordination of the experimental design or execution between the two laboratories. Indeed, the two laboratories were not aware of each other's experiments until the studies were completed. Male and female APPPS1-21 mice were treated daily with 40, 80, or 160 mg/kg of GV-971 from 8, when Aß burden was detectable upto 12 weeks of age when Aß burden was near maximal levels. In parallel, and to corroborate existing published studies and further investigate sex-related differences, male and female 5XFAD mice were treated daily with 100 mg/kg of GV-971 from 7 to 9 months of age when Aß burden was near peak levels. Subsequently, the two laboratories independently assessed amyloid-ß deposition, metagenomic, and neuroinflammatory profiles. Finally, studies were initiated at the University of Chicago to evaluate the metabolites in cecal tissue from vehicle and GV-971-treated 5XFAD mice.Results These studies showed that independent of the procedural differences (dosage, timing and duration of treatment) between the two laboratories, cerebral amyloidosis was reduced primarily in male mice, independent of strain. We also observed sex-specific microbiota differences following GV-971 treatment. Interestingly, GV-971 significantly altered multiple overlapping bacterial species at both institutions. Moreover, we discovered that GV-971 significantly impacted microbiome metabolism, particularly by elevating amino acid production and influencing the tryptophan pathway. The metagenomics and metabolomics changes correspond with notable reductions in peripheral pro-inflammatory cytokine and chemokine profiles. Furthermore, GV-971 treatment dampened astrocyte and microglia activation, significantly decreasing plaque-associated reactive microglia while concurrently increasing homeostatic microglia only in male mice. Bulk RNAseq analysis unveiled sex-specific changes in cerebral cortex transcriptome profiles, but most importantly, the transcriptome changes in the GV-971-treated male group revealed the involvement of microglia and inflammatory responses.Conclusions In conclusion, these studies demonstrate the connection between the gut microbiome, neuroinflammation, and Alzheimer's disease pathology while highlighting the potential therapeutic effect of GV-971. GV-971 targets the microbiota-microglia-amyloid axis, leading to the lowering of plaque pathology and neuroinflammatory signatures in a sex-dependent manner when given at the onset of Aß deposition or when given after Aß deposition is already at higher levels.

Amelioration of Tau and ApoE4-linked glial lipid accumulation and neurodegeneration with an LXR agonist.

Apolipoprotein E (APOE) is a strong genetic risk factor for late-onset Alzheimer's disease (LOAD). APOE4 increases and APOE2 decreases risk relative to APOE3. In the P301S mouse model of tauopathy, ApoE4 increases tau pathology and neurodegeneration when compared with ApoE3 or the absence of ApoE. However, the role of ApoE isoforms and lipid metabolism in contributing to tau-mediated degeneration is unknown. We demonstrate that in P301S tau mice, ApoE4 strongly promotes glial lipid accumulation and perturbations in cholesterol metabolism and lysosomal function. Increasing lipid efflux in glia via an LXR agonist or Abca1 overexpression strongly attenuates tau pathology and neurodegeneration in P301S/ApoE4 mice. We also demonstrate reductions in reactive astrocytes and microglia, as well as changes in cholesterol biosynthesis and metabolism in glia of tauopathy mice in response to LXR activation. These data suggest that promoting efflux of glial lipids may serve as a therapeutic approach to ameliorate tau and ApoE4-linked neurodegeneration.

Parenchymal border macrophages regulate tau pathology and tau-mediated neurodegeneration.

Parenchymal border macrophages (PBMs) reside close to the central nervous system parenchyma and regulate CSF flow dynamics. We recently demonstrated that PBMs provide a clearance pathway for amyloid-ß peptide, which accumulates in the brain in Alzheimer's disease (AD). Given the emerging role for PBMs in AD, we explored how tau pathology affects the CSF flow and the PBM populations in the PS19 mouse model of tau pathology. We demonstrated a reduction of CSF flow, and an increase in an MHCII+PBM subpopulation in PS19 mice compared with WT littermates. Consequently, we asked whether PBM dysfunction could exacerbate tau pathology and tau-mediated neurodegeneration. Pharmacological depletion of PBMs in PS19 mice led to an increase in tau pathology and tau-dependent neurodegeneration, which was independent of gliosis or aquaporin-4 depolarization, essential for the CSF-ISF exchange. Together, our results identify PBMs as novel cellular regulators of tau pathology and tau-mediated neurodegeneration.

APOE-ε4 synergizes with sleep disruption to accelerate Aß deposition and Aß-associated tau seeding and spreading.

Alzheimer's disease (AD) is the most common cause of dementia. The APOE-ε4 allele of the apolipoprotein E (APOE) gene is the strongest genetic risk factor for late-onset AD. The APOE genotype modulates the effect of sleep disruption on AD risk, suggesting a possible link between apoE and sleep in AD pathogenesis, which is relatively unexplored. We hypothesized that apoE modifies Aß deposition and Aß plaque-associated tau seeding and spreading in the form of neuritic plaque-tau (NP-tau) pathology in response to chronic sleep deprivation (SD) in an apoE isoform-dependent fashion. To test this hypothesis, we used APPPS1 mice expressing human APOE-ε3 or -ε4 with or without AD-tau injection. We found that SD in APPPS1 mice significantly increased Aß deposition and peri-plaque NP-tau pathology in the presence of APOE4 but not APOE3. SD in APPPS1 mice significantly decreased microglial clustering around plaques and aquaporin-4 (AQP4) polarization around blood vessels in the presence of APOE4 but not APOE3. We also found that sleep-deprived APPPS1:E4 mice injected with AD-tau had significantly altered sleep behaviors compared with APPPS1:E3 mice. These findings suggest that the APOE-ε4 genotype is a critical modifier in the development of AD pathology in response to SD.

Sleep deprivation exacerbates microglial reactivity and Aß deposition in a TREM2-dependent manner in mice.

Sleep loss is associated with cognitive decline in the aging population and is a risk factor for Alzheimer's disease (AD). Considering the crucial role of immunomodulating genes such as that encoding the triggering receptor expressed on myeloid cells type 2 (TREM2) in removing pathogenic amyloid-ß (Aß) plaques and regulating neurodegeneration in the brain, our aim was to investigate whether and how sleep loss influences microglial function in mice. We chronically sleep-deprived wild-type mice and the 5xFAD mouse model of cerebral amyloidosis, expressing either the humanized TREM2 common variant, the loss-of-function R47H AD-associated risk variant, or without TREM2 expression. Sleep deprivation not only enhanced TREM2-dependent Aß plaque deposition compared with 5xFAD mice with normal sleeping patterns but also induced microglial reactivity that was independent of the presence of parenchymal Aß plaques. We investigated lysosomal morphology using transmission electron microscopy and found abnormalities particularly in mice without Aß plaques and also observed lysosomal maturation impairments in a TREM2-dependent manner in both microglia and neurons, suggesting that changes in sleep modified neuro-immune cross-talk. Unbiased transcriptome and proteome profiling provided mechanistic insights into functional pathways triggered by sleep deprivation that were unique to TREM2 and Aß pathology and that converged on metabolic dyshomeostasis. Our findings highlight that sleep deprivation directly affects microglial reactivity, for which TREM2 is required, by altering the metabolic ability to cope with the energy demands of prolonged wakefulness, leading to further Aß deposition, and underlines the importance of sleep modulation as a promising future therapeutic approach.

Astrocytic APOE4 removal confers cerebrovascular protection despite increased cerebral amyloid angiopathy.

BACKGROUND: Alzheimer Disease (AD) and cerebral amyloid angiopathy (CAA) are both characterized by amyloid-ß (Aß) accumulation in the brain, although Aß deposits mostly in the brain parenchyma in AD and in the cerebrovasculature in CAA. The presence of CAA can exacerbate clinical outcomes of AD patients by promoting spontaneous intracerebral hemorrhage and ischemia leading to CAA-associated cognitive decline. Genetically, AD and CAA share the ε4 allele of the apolipoprotein E (APOE) gene as the strongest genetic risk factor. Although tremendous efforts have focused on uncovering the role of APOE4 on parenchymal plaque pathogenesis in AD, mechanistic studies investigating the role of APOE4 on CAA are still lacking. Here, we addressed whether abolishing APOE4 generated by astrocytes, the major producers of APOE, is sufficient to ameliorate CAA and CAA-associated vessel damage. METHODS: We generated transgenic mice that deposited both CAA and plaques in which APOE4 expression can be selectively suppressed in astrocytes. At 2-months-of-age, a timepoint preceding CAA and plaque formation, APOE4 was removed from astrocytes of 5XFAD APOE4 knock-in mice. Mice were assessed at 10-months-of-age for Aß plaque and CAA pathology, gliosis, and vascular integrity. RESULTS: Reducing the levels of APOE4 in astrocytes shifted the deposition of fibrillar Aß from the brain parenchyma to the cerebrovasculature. However, despite increased CAA, astrocytic APOE4 removal reduced overall Aß-mediated gliosis and also led to increased cerebrovascular integrity and function in vessels containing CAA. CONCLUSION: In a mouse model of CAA, the reduction of  APOE4 derived specifically from astrocytes, despite increased fibrillar Aß deposition in the vasculature, is sufficient to reduce Aß-mediated gliosis and cerebrovascular dysfunction.

TREM2-independent microgliosis promotes tau-mediated neurodegeneration in the presence of ApoE4.

In addition to tau and Aß pathologies, inflammation plays an important role in Alzheimer's disease (AD). Variants in APOE and TREM2 increase AD risk. ApoE4 exacerbates tau-linked neurodegeneration and inflammation in P301S tau mice and removal of microglia blocks tau-dependent neurodegeneration. Microglia adopt a heterogeneous population of transcriptomic states in response to pathology, at least some of which are dependent on TREM2. Previously, we reported that knockout (KO) of TREM2 attenuated neurodegeneration in P301S mice that express mouse Apoe. Because of the possible common pathway of ApoE and TREM2 in AD, we tested whether TREM2 KO (T2KO) would block neurodegeneration in P301S Tau mice expressing ApoE4 (TE4), similar to that observed with microglial depletion. Surprisingly, we observed exacerbated neurodegeneration and tau pathology in TE4-T2KO versus TE4 mice, despite decreased TREM2-dependent microgliosis. Our results suggest that tau pathology-dependent microgliosis, that is, TREM2-independent microgliosis, facilitates tau-mediated neurodegeneration in the presence of ApoE4.

Role of anterior cingulate cortex inputs to periaqueductal gray for pain avoidance.

Although pain-related excessive fear is known to be a key factor in chronic pain disability, which involves the anterior cingulate cortex (ACC), little is known about the downstream circuits of the ACC for fear avoidance in pain processing. Using behavioral experiments and functional magnetic resonance imaging with optogenetics at 15.2 T, we demonstrate that the ACC is a part of the abnormal circuit changes in chronic pain and its downstream circuits are closely related to modulating sensorimotor integration and generating active movement rather than carrying sensory information. The projection from the ACC to the dorsolateral and lateral parts of the periaqueductal gray (dl/lPAG) especially enhances both reflexive and active avoidance behavior toward pain. Collectively, our results indicate that increased signals from the ACC to the dl/lPAG might be critical for excessive fear avoidance in chronic pain disability.

Laparoscopic cholecystectomy in a swine model using a novel near-infrared fluorescent IV dye (BL-760).

BACKGROUND AND OBJECTIVES: Bile duct injury during laparoscopic cholecystectomy has an incidence rate of 1%-2% and commonly appears under conditions of severe inflammation, adhesion, or unexpected anatomical variations. Despite the difficulties and rising concerns of identifying bile duct during surgeries, surgeons do not have a specific modality to identify bile duct except intraoperative cholangiography. While no biliary-specific fluorescent dye exists for clinical use, our team has previously described the development of a preclinical biliary-specific dye, BL-760. Here, we present our study of laparoscopic cholecystectomy using the fluorescent dye in a swine model. STUDY DESIGN/MATERIALS AND METHODS: With an approval from Institutional Animal Care and Use Committee, two 20-25 kg swine underwent laparoscopic abdominal surgery using a Food and Drug Administration-cleared fluorescent laparoscopic system. Images of the liver and gallbladder were taken both before and after intravenous injection of the novel fluorescent dye. The dye was dosed at 60 µg/kg and injected via the ear vein. The amount of time taken to visualize fluorescence in the biliary tract was measured. Fluorescent signal was observed after injection, and target-to-background ratio (TBR) of the biliary tract to surrounding cystic artery and liver parenchyma was measured. RESULTS: Biliary tract visualization under fluorescent laparoscopy was achieved within 5 min after the dye injection without any adverse effects. Cystic duct and extrahepatic duct were clearly visualized and identified with TBR values of 2.19 and 2.32, respectively, whereas no fluorescent signal was detected in liver. Cystic duct and artery were successfully ligated by an endoscopic clip applier with the visual assistance of highlighted biliary tract images. Laparoscopic cholecystectomy was completed within 30 min in each case without any complications. CONCLUSIONS: BL-760 is a novel preclinical fluorescent dye useful for intraoperative identification and visualization of biliary tract. Such fluorescent dye that is exclusively metabolized by liver and rapidly excreted into biliary tract would be beneficial for all types of hepato-biliary surgeries. With the validation of additional preclinical data, this novel dye has potential to be a valuable tool to prevent any iatrogenic biliary injuries and/or bile leaks during laparoscopic abdominal and liver surgeries.

A Novel Multidrug Combination Mitigates Rat Liver Steatosis Through Activating AMPK Pathway During Normothermic Machine Perfusion.

BACKGROUND: Hepatic steatosis is now the leading cause of liver discards in deceased donors. Previous studies [Yarmush formula (Y) defatting] have successfully reduced the fat content by treating rat steatotic livers on extracorporeal normothermic machine perfusion (NMP) with a multidrug combination including the GW compounds that were linked to an increased risk of carcinogenesis. METHODS: We developed a novel multidrug combination by replacing the GW compounds with 2 polyphenols, epigallocatechin-3-gallate (E) and resveratrol (R). Sixteen rat livers were placed on NMP and assigned to control, Y defatting, Y + E + R defatting, or Y'-GW + E + R defatting groups (Y'-GW = 90% dose-reduced Y defatting, n = 4/group). RESULTS: All livers in defatting groups had significant decreases in hepatic triglyceride content at the end of the experiment. However, livers treated with our novel Y'-GW + E + R combination had evidence of increased metabolism and less hepatocyte damage and carcinogenic potential. Our Y'-GW + E + R combination had increased phosphorylation of AMP-activated protein kinase (P = 0.019) and acetyl-CoA carboxylase (P = 0.023) compared with control; these were not increased in Y + E + R group and actually decreased in the Y group. Furthermore, the Y'-GW + E + R group had less evidence of carcinogenic potential with no increase in AKT phosphorylation compared with control (P = 0.089); the Y (P = 0.031) and Y + E + R (P = 0.035) groups had striking increases in AKT phosphorylation. Finally, our Y'-GW + E + R showed less evidence of hepatocyte damage with significantly lower perfusate alanine aminotransferase (P = 0.007) and aspartate aminotransferase (P = 0.014) levels. CONCLUSIONS: We have developed a novel multidrug combination demonstrating promising defatting efficacy via activation of the AMP-activated protein kinase pathway with an optimized safety profile and reduced hepatotoxicity during ex vivo NMP.

Goal-directed interaction of stimulus and task demand in the parahippocampal region.

The hippocampus and parahippocampal region are essential for representing episodic memories involving various spatial locations and objects, and for using those memories for future adaptive behavior. The "dual-stream model" was initially formulated based on anatomical characteristics of the medial temporal lobe, dividing the parahippocampal region into two streams that separately process and relay spatial and nonspatial information to the hippocampus. Despite its significance, the dual-stream model in its original form cannot explain recent experimental results, and many researchers have recognized the need for a modification of the model. Here, we argue that dividing the parahippocampal region into spatial and nonspatial streams a priori may be too simplistic, particularly in light of ambiguous situations in which a sensory cue alone (e.g., visual scene) may not allow such a definitive categorization. Upon reviewing evidence, including our own, that reveals the importance of goal-directed behavioral responses in determining the relative involvement of the parahippocampal processing streams, we propose the Goal-directed Interaction of Stimulus and Task-demand (GIST) model. In the GIST model, input stimuli such as visual scenes and objects are first processed by both the postrhinal and perirhinal cortices-the postrhinal cortex more heavily involved with visual scenes and perirhinal cortex with objects-with relatively little dependence on behavioral task demand. However, once perceptual ambiguities are resolved and the scenes and objects are identified and recognized, the information is then processed through the medial or lateral entorhinal cortex, depending on whether it is used to fulfill navigational or non-navigational goals, respectively. As complex sensory stimuli are utilized for both navigational and non-navigational purposes in an intermixed fashion in naturalistic settings, the hippocampus may be required to then put together these experiences into a coherent map to allow flexible cognitive operations for adaptive behavior to occur.

Impairment of Pattern Separation of Ambiguous Scenes by Single Units in the CA3 in the Absence of the Dentate Gyrus.

Theoretical models and experimental evidence have suggested that connections from the dentate gyrus (DG) to CA3 play important roles in representing orthogonal information (i.e., pattern separation) in the hippocampus. However, the effects of eliminating the DG on neural firing patterns in the CA3 have rarely been tested in a goal-directed memory task that requires both the DG and CA3. In this study, selective lesions in the DG were made using colchicine in male Long-Evans rats, and single units from the CA3 were recorded as the rats performed visual scene memory tasks. The original scenes used in training were altered during testing by blurring to varying degrees or by using visual masks, resulting in maximal recruitment of the DG-CA3 circuits. Compared with controls, the performance of rats with DG lesions was particularly impaired when blurred scenes were used in the task. In addition, the firing rate modulation associated with visual scenes in these rats was significantly reduced in the single units recorded from the CA3 when ambiguous scenes were presented, largely because DG-deprived CA3 cells did not show stepwise, categorical rate changes across varying degrees of scene ambiguity compared with controls. These findings suggest that the DG plays key roles not only during the acquisition of scene memories but also during retrieval when modified visual scenes are processed in conjunction with the CA3 by making the CA3 network respond orthogonally to ambiguous scenes.SIGNIFICANCE STATEMENT Despite the behavioral evidence supporting the role of the dentate gyrus in pattern separation in the hippocampus, the underlying neural mechanisms are largely unknown. By recording single units from the CA3 in DG-lesioned rats performing a visual scene memory task, we report that the scene-related modulation of neural firing was significantly reduced in the DG-lesion rats compared with controls, especially when the original scene stimuli were ambiguously altered. Our findings suggest that the dentate gyrus plays an essential role during memory retrieval and performs a critical computation to make categorical rate modulation occur in the CA3 between different scenes, especially when ambiguity is present in the environment.

A Biliary Tract-Specific Near-Infrared Fluorescent Dye for Image-Guided Hepatobiliary Surgery.

Despite advances, visual inspection, palpation, and intraoperative ultrasound remain the most utilized tools during surgery today. A particularly challenging issue is the identification of the biliary system due to its complex architecture partially embedded within the liver. Fluorescence guided surgical interventions, particularly using near-infrared (NIR) wavelengths, are an emerging approach for the real-time assessment of the hepatobiliary system. However, existing fluorophores, such as the FDA-approved indocyanine green (ICG), have significant limitations for rapid and selective visualization of bile duct anatomy. Here we report a novel NIR fluorophore, BL (Bile Label)-760, which is exclusively metabolized by the liver providing high signal in the biliary system shortly after intravenous administration. This molecule was identified by first screening a small set of known heptamethine cyanines including clinically utilized agents. After finding that none of these were well-suited, we then designed and tested a small series of novel dyes within a prescribed polarity range. We validated the molecule that emerged from these efforts, BL-760, through animal studies using both rodent and swine models employing a clinically applicable imaging system. In contrast to ICG, BL-760 fluorescence revealed a high target-to-background ratio (TBR) of the cystic duct relative to liver parenchyma 5 min after intravenous injection. During hepatic resection surgery, intrahepatic ducts were clearly highlighted, and bile leakage was easily detected. In conclusion, BL-760 has highly promising properties for intraoperative navigation during hepatobiliary surgery.

High Prevalence of Cefotaxime Resistant Bacteria in Grazing Beef Cattle: A Cross Sectional Study.

Although the over-use of antibiotics during food animal production is a potential driver of antimicrobial resistant microorganisms (ARMs), a high prevalence of cefotaxime resistant bacteria (CRB) has been observed in grazing animals raised without antibiotic supplementation. In this cross-sectional study, the prevalence and concentration of CRB in beef cattle on grazing farms were investigated. Fecal samples from the recto-anal junction of cattle (n = 840) and environmental samples (n = 258) were collected from 17 farms in North and Central Florida in the United States, and a survey of farm characteristics, animal husbandry practices, and antibiotic usage was conducted. CRB were detected in fecal samples from 47.4% of all cattle, with the prevalence ranging from 21.1 to 87.5% on farms, and significantly higher (P < 0.001) in calves compared to adult cows (54.1 vs. 41.8%). Environmental samples had a higher prevalence than fecal samples (P < 0.001), with CRB detected in 88.6% of water, 98.7% of soil, and 95.7% of forage samples. Compared to the concentration (log CFU/g) of CRB in fecal samples (2.95, 95% CI: 2.89, 3.02), the concentration of CRB was higher (P < 0.001) in soil and forage samples (5.37, 95% CI: 5.16, 5.57) and lower (P < 0.001) in water samples (1.08, 95% CI: 0.82, 1.36). Soil microbiota from farms with high prevalence of CRB clustered closer together and the proportion of Phylum Proteobacteria was higher on farms with high prevalence of CRB resistance. Large farming operations were associated with a 58% higher likelihood of CRB detection in fecal samples. Regular cleaning of drinking troughs and the addition of ionophores to feed were associated with CRB reduction in fecal samples. Taken together, the widespread of CRB into both cattle seldom treated with cephalosporin antibiotics and the surrounding environment suggests the environment is a natural source of antimicrobial resistance in beef cattle.

Comprehensive in vitro and in vivo risk assessments of chitosan microparticles using human epithelial cells and Caenorhabditis elegans.

The safety of using nano- and microparticles is a developing concern. In this study, we conducted risk assessments of chitosan microparticles (CMs) using in vitro human epithelial cell lines and in vivo animal model, Caenorhabditis elegans. After engineering of various CMs, we screened four CMs based on antimicrobial activity, which is a potential usage for disease treatment caused by multidrug resistant bacteria, and evaluated for risk assessments. CMs, with strong antimicrobial activity, and inorganic nanoparticles (SiO2, TiO2, and ZnO) did not cause toxicity in human cells measured by cell membrane integrity, mitochondria activity, and reactive oxygen species concentration. However, when applied to C. elegans, only CMs generated with low molecular weight chitosan and tripolyphosphate at 0.1% did not affect the lifespan, while the other CMs and inorganic nanoparticles shortened the lifespan, suggesting that they may cause subtle toxicity. These results suggest that C. elegans could be a sensitive animal model to measure low level of toxicity of nano- and microparticles. Taken together, although CMs do not cause toxicity at working concentrations of antimicrobial activity in human epithelial cells, they may cause toxicity at high concentration, suggesting that nano- and microparicles should be thoroughly investigated before they are applied.

Cartilage Regeneration in Osteoarthritic Patients by a Composite of Allogeneic Umbilical Cord Blood-Derived Mesenchymal Stem Cells and Hyaluronate Hydrogel: Results from a Clinical Trial for Safety and Proof-of-Concept with 7 Years of Extended Follow-Up.

Few methods are available to regenerate articular cartilage defects in patients with osteoarthritis. We aimed to assess the safety and efficacy of articular cartilage regeneration by a novel medicinal product composed of allogeneic human umbilical cord blood-derived mesenchymal stem cells (hUCB-MSCs). Patients with Kellgren-Lawrence grade 3 osteoarthritis and International Cartilage Repair Society (ICRS) grade 4 cartilage defects were enrolled in this clinical trial. The stem cell-based medicinal product (a composite of culture-expanded allogeneic hUCB-MSCs and hyaluronic acid hydrogel [Cartistem]) was applied to the lesion site. Safety was assessed by the World Health Organization common toxicity criteria. The primary efficacy outcome was ICRS cartilage repair assessed by arthroscopy at 12 weeks. The secondary efficacy outcome was visual analog scale (VAS) score for pain on walking. During a 7-year extended follow-up, we evaluated safety, VAS score, International Knee Documentation Committee (IKDC) subjective score, magnetic resonance imaging (MRI) findings, and histological evaluations. Seven participants were enrolled. Maturing repair tissue was observed at the 12-week arthroscopic evaluation. The VAS and IKDC scores were improved at 24 weeks. The improved clinical outcomes were stable over 7 years of follow-up. The histological findings at 1 year showed hyaline-like cartilage. MRI at 3 years showed persistence of the regenerated cartilage. Only five mild to moderate treatment-emergent adverse events were observed. There were no cases of osteogenesis or tumorigenesis over 7 years. The application of this novel stem cell-based medicinal product appears to be safe and effective for the regeneration of durable articular cartilage in osteoarthritic knees. Stem Cells Translational Medicine 2017;6:613-621.

Restoration of a large osteochondral defect of the knee using a composite of umbilical cord blood-derived mesenchymal stem cells and hyaluronic acid hydrogel: a case report with a 5-year follow-up.

BACKGROUND: The treatment of articular cartilage defects is a therapeutic challenge for orthopaedic surgeons. Furthermore, large osteochondral defects needs restoration of the underlying bone for sufficient biomechanical characteristics as well as the overlying cartilage. CASE PRESENTATION: A symptomatic large osteochondral defect in the knee joint was restored using a composite of umbilical cord blood-derived mesenchymal stem cells (UCB-MSCs) 0.5 x 107/ml and 4% hyaluronic acid (HA) hydrogel. Significant improvements in pain and function of the knee joint were identified by the evaluation at 12 months after surgery. A hyaline-like cartilage completely filled the defect and was congruent with the surrounding normal cartilage as revealed by magnetic resonance imaging (MRI), a second-look arthroscopy and histological assessment. The improved clinical outcomes maintained until 5.5 years. MRI also showed the maintenance of the restored bony and cartilaginous tissues. CONCLUSION: This case report suggests that the composite of allogeneic UCB-MSCs and HA hydrogel can be considered a safe and effective treatment option for large osteochondral defects of the knee.

Dual-Channel Endoscopic Indocyanine Green Fluorescence Angiography for Clipping of Cerebral Aneurysms.

OBJECTIVE: Neuroendoscopy is useful for assessing status of perforators, parent arteries, and aneurysms beyond the straight line of microscopic view during aneurysm clipping. We aimed to evaluate the clinical usefulness of our endoscopic indocyanine green angiography (ICGA) system, which can simultaneously display visible light and indocyanine green fluorescent images. METHODS: Surgical clipping of 16 unruptured aneurysms in 10 patients was performed via the keyhole approach. Using our endoscopic ICGA and commercial microscopic ICGA systems, we prospectively compared 10 targeted cerebral aneurysms at the posterior communicating (n = 4) and anterior choroidal (n = 6) arteries. RESULTS: Microscopic ICGA and endoscopic ICGA were feasible during surgery. Microscopic ICGA displayed 50% of branch orifices, 100% of branch trunks, and 20% of exact clip positions, whereas endoscopic ICGA showed 100% of these. Based on endoscopic ICGA findings such as incomplete clipping and compromise of parent arteries or branches, clips were repositioned in 2 cases, and additional clips were applied in 2 cases. Complete occlusion and residual neck states were achieved in 6 and 4 aneurysms after surgery. There were no neurologic deficits within 3 months after surgery except for frontalis palsy and anosmia in each patient. CONCLUSIONS: The endoscopic ICGA system with dual imaging of visible light and indocyanine green fluorescence was very useful for assessing geometry of aneurysms and surrounding vessels before clipping and for evaluating completeness of clip position after clipping.

Antioxidant Activity of Brown Soybean Ethanolic Extracts and Application to Cooked Pork Patties.

The brown soybean extract (BE, extracted by distilled water, 50%, 75%, and 95% ethanol) were analyzed for their total phenol, flavonoid, anthocyanin content, and DPPH radical-scavenging activity to determine antioxidant activities. Brown soybean extract with 75% ethanol showed significantly higher DPPH radical scavenging activity, total phenol and anthocyanin content compared to the other treatments (p<0.05). Then, brown soybean extract with 75% ethanol was applied to pork patties at different concentration (0.05%, 0.1%, and 0.2%) and lipid oxidation was evaluated during 15 d of refrigerated storage. Addition of BE significantly increased redness and pH values, respectively (p<0.05). Moreover, TBARS value of pork patties decreased significantly (p<0.05) as BE concentration increased. In sensory evaluation, pork patties with 0.1% BE had significantly higher score than other treatments in flavor and overall acceptability (p<0.05). Consequently, these results indicate that 0.1% BE could be an effective natural antioxidant to inhibit lipid oxidation in pork patties.