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In Alzheimer's disease, amyloid deposits along the brain vasculature lead to a condition known as cerebral amyloid angiopathy (CAA), which impairs blood-brain barrier (BBB) function and accelerates cognitive degeneration. Apolipoprotein (APOE4) is the strongest risk factor for CAA, yet the mechanisms underlying this genetic susceptibility are unknown. Here we developed an induced pluripotent stem cell-based three-dimensional model that recapitulates anatomical and physiological properties of the human BBB in vitro. Similarly to CAA, our in vitro BBB displayed significantly more amyloid accumulation in APOE4 compared to APOE3. Combinatorial experiments revealed that dysregulation of calcineurin-nuclear factor of activated T cells (NFAT) signaling and APOE in pericyte-like mural cells induces APOE4-associated CAA pathology. In the human brain, APOE and NFAT are selectively dysregulated in pericytes of APOE4 carriers, and inhibition of calcineurin-NFAT signaling reduces APOE4-associated CAA pathology in vitro and in vivo. Our study reveals the role of pericytes in APOE4-mediated CAA and highlights calcineurin-NFAT signaling as a therapeutic target in CAA and Alzheimer's disease.

The purpose of this study was to demonstrate the effectiveness of controlled telescoping system for lateral hip pain caused by sliding of the blade following intramedullary nailing of trochanteric fractures.A retrospective cohort study was performed to compare the controlled telescoping system (Compression Hip Nail; CHN) with the conventional sliding system (Proximal Femoral Nail Antirotation; PFNA) for trochanteric fractures. 74 cases in the PFNA group and 77 cases in the CHN group were included from two university hospitals in this study. All patients had a minimum of 12-month follow up period. Lateral hip pain was evaluated and operation time and blood loss during the surgery were measured. The fracture classification was evaluated. The quality of postoperative reduction and other complications after surgery were also evaluated and tip-apex distance (TAD), telescoping and lateral protrusion of the blade and lag screw were measured.The mean age was 78.5 years in the PFNA group and 74.7 years in the CHN group (p=0.25). The mean telescoping was 19.2mm in the PFNA group and 10.7mm in the CHN group (p0.001). The mean length of lateral protrusion was 10.5mm in the PFNA group and 2.5mm in the CHN group (p0.001). Twenty-eight patients in the PFNA group complained of lateral hip pain, whereas 12 patients in the CHN group did (p=0.002). These four variables showed statistically significant differences between the PFNA and CHN groups (p0.05). The length of lateral protrusion was the only variable significantly related to lateral hip pain through multivariate logistic regression analysis (p=0.045).The degree of lateral protrusion was mainly related to lateral hip pain. Therefore, controlled telescoping would help to decrease lateral hip pain by decreasing the lateral protrusion beyond the lateral femoral cortex.

Increased p25, a proteolytic fragment of the regulatory subunit p35, is known to induce aberrant activity of cyclin-dependent kinase 5 (Cdk5), which is associated with neurodegenerative disorders, including Alzheimer's disease. Previously, we showed that replacing endogenous p35 with the noncleavable mutant p35 (Δp35) attenuated amyloidosis and improved cognitive function in a familial Alzheimer's disease mouse model. Here, to address the role of p25/Cdk5 in tauopathy, we generated double-transgenic mice by crossing mice overexpressing mutant human tau (P301S) with Δp35KImice. We observed significant reduction of phosphorylated tau and its seeding activity in the brain of double transgenic mice compared with the P301S mice. Furthermore, synaptic loss and impaired LTP at hippocampal CA3 region ofP301Smice were attenuated by blocking p25 generation. To further validate the role of p25/Cdk5 in tauopathy, we used frontotemporal dementia patient-derived induced pluripotent stem cells (iPSCs) carrying the Tau P301L mutation and generated P301L:Δp35KI isogenic iPSC lines using CRISPR/Cas9 genome editing. We created cerebral organoids from the isogenic iPSCs and found that blockade of p25 generation reduced levels of phosphorylated tau and increased expression of synaptophysin. Together, these data demonstrate a crucial role for p25/Cdk5 in mediating tau-associated pathology and suggest that inhibition of this kinase can remedy neurodegenerative processes in the presence of pathogenic tau mutation.SIGNIFICANCE STATEMENTAccumulation of p25 results in aberrant Cdk5 activation and induction of numerous pathological phenotypes, such as neuroinflammation, synaptic loss, Aβ accumulation, and tau hyperphosphorylation. However, it was not clear whether p25/Cdk5 activity is necessary for the progression of these pathological changes. We recently developed the Δp35KItransgenic mouse that is deficient in p25 generation and Cdk5 hyperactivation. In this study, we used this mouse model to elucidate the role of p25/Cdk5 in FTD mutant tau-mediated pathology. We also used a frontotemporal dementia patient-derived induced pluripotent stem cell carrying the Tau P301L mutation and generated isogenic lines in which p35 is replaced with noncleavable mutant Δp35. Our data suggest that p25/Cdk5 plays an important role in tauopathy in both mouse and human model systems.

Induced pluripotent stem cells (iPSCs) have radically advanced the field of regenerative medicine by making possible the production of patient-specific pluripotent stem cells from adult individuals. While cell differentiation protocols have been successfully developed, and animal models of human disease have proved that these cells have the potential to treat human diseases and conditions produced as a consequence of aging, degeneration, injury, and birth defects, logistical issues still remain unsolved and hamper the possibility of testing these cells in human clinical trials. Among them is the widely spread use of animal products for the generation and culture of iPSCs. We report here a xeno-free iPSC generation system that addresses all the steps of iPSCs production including the isolation and culture of adult skin fibroblasts, and iPSCs generation, expansion, and maintenance. iPSCs generated with a polycistronic lentiviral vector under xeno-free conditions displayed markers of pluripotency and gave rise to embryoid bodies (EBs) displaying indicators of the 3 primary germ layers. Xeno-free iPSCs injected into nude mice produced classic teratomas, and teratoma explants cultured under conditions favoring fibroblastic cells gave rise to cells morphologically indistinguishable from input cells. Protocols here described will facilitate the implementation of new cellular therapies for preclinical and clinical studies, potentially reducing the regulatory burden without compromising the differentiation potential of the cells.

The apolipoprotein E4 (APOE4) variant is the single greatest genetic risk factor for sporadic Alzheimer's disease (sAD). However, the cell-type-specific functions of APOE4 in relation to AD pathology remain understudied. Here, we utilize CRISPR/Cas9 and induced pluripotent stem cells (iPSCs) to examine APOE4 effects on human brain cell types. Transcriptional profiling identified hundreds of differentially expressed genes in each cell type, with the most affected involving synaptic function (neurons), lipid metabolism (astrocytes), and immune response (microglia-like cells). APOE4 neurons exhibited increased synapse number and elevated Aβ42 secretion relative to isogenic APOE3 cells while APOE4 astrocytes displayed impaired Aβ uptake and cholesterol accumulation. Notably, APOE4 microglia-like cells exhibited altered morphologies, which correlated with reduced Aβ phagocytosis. Consistently, converting APOE4 to APOE3 in brain cell types from sAD iPSCs was sufficient to attenuate multiple AD-related pathologies. Our study establishes a reference for human cell-type-specific changes associated with the APOE4 variant. Video Abstract: [Figure presented] By generating and characterizing isogenic APOE3- or APOE4-carrying human brain cell types, Lin et al. show that the APOE4 variant can lead to extensive gene expression alterations, and multiple cellular phenotypes potentially related to AD pathogenesis, in neurons, astrocytes, and microglia., National Institutes of Health (NIH) (Grants RF1-AG048056, RC1-AG036106, and RF1-AG048029)

Fluorescent live imaging of cells and embryos at subcellular resolution poses significant challenges for biologists due to morbidity and mortality ensuing from phototoxicity. Here we report the use of a spinning-disk confocal microscope to image mouse and bovine preimplantation embryos without impairing their developmental potential. We also present data indicating that this imaging technique does not affect the functionality of subcellular components as assessed by reactive oxygen species (ROS) production, caspase activity, and DNA integrity. Spinning-disk confocal microscopy was also useful in determining cell number and allocation in transgenic bovine blastocysts. We conclude that this imaging method is suitable for monitoring preimplantation embryos.

The transport mechanism mediating brain uptake of tumor necrosis factor (TNF)-α has been studied. When125I-labeled rat TNF-α was used in internal carotid artery perfusions in rats, the cytokine showed transcytosis through the blood-brain barrier in intact form (permeability-surface area product 0.34 ± 0.13 μl · min−1· g−1). Uptake was inhibited by low nanomolar concentrations of unlabeled rat TNF-α. Human TNF-α, which does not interact with the p80 TNF receptor in rodents, showed no brain uptake. mRNA expression of both p60 and p80 receptors could be demonstrated in native brain microvessel preparations. These transcripts increased to 149% (p60) and 127% (p80) of control 4 h after a systemic immune stimulation (2 mg/kg bacterial endotoxin ip). Lipopolysaccharide treatment did not alter the rate of brain uptake of TNF-α measured between 4 and 24 h later. In conclusion, a receptor-mediated mechanism is responsible for the transcytosis of TNF-α. Saturable transport, requiring the p80 receptor, occurs at concentrations encountered under pathophysiological conditions and therefore constitutes a relevant mechanism of communication between the immune system and the brain.

The purpose of the present retrospective study was to describe the single oblique posterolateral approach for open reduction and internal fixation of large, displaced, posterior malleolar fractures with an associated lateral malleolar fracture. A single oblique posterolateral approach was used for osteosynthesis of the posterior and lateral malleolus in 50 consecutive patients (23 females [46%], 27 males [54%]; mean age, 47.44 ± 16.13 years; mean follow-up duration, 26.32 ± 5.15 months). The mean interval to surgery was 4.3 ± 1.9 days after the inciting trauma. During the follow-up period, the surgery was complicated by skin necrosis around the incision in 2 (4%) patients and sural nerve damage in 2 (4%) patients. We found that the single oblique posterolateral approach to large, displaced, posterior malleolar fractures with an associated lateral malleolar fracture provided easy exposure of the posterior and lateral malleoli and had the potential to decrease the incidence of sural nerve injury because of the smaller incision size.

Previous reports have shown that antidepressants increase neuronal cell proliferation and enhance neuroplasticity both in vivo and in vitro. This study investigated the direct effects of one such antidepressant, fluoxetine , on cell proliferation and on the production of neurotrophic factors in neuronal precursors derived from human embryonic stem cells (hESCs; H9). Fluoxetine induced the differentiation of neuronal precursors, strongly enhancing neuronal characteristics. The rate of proliferation was higher in fluoxetine -treated cells than in control cells, as determined by MTT [3(4,5-dimethylthiazol-2-yl) 2,5-diphenyltetrazolium bromide] assay. The CPDL (cumulative population doubling level) of the fluoxetine-treated cells was significantly increased in comparison to that of control cells (p.001). Bromodeoxyuridine incorporation and staurosporine-induced apoptosis assays were elevated in fluoxetine-treated cells. Quantitative RT-PCR analysis revealed no significant differences in the expression of neurotrophic factors, brain-derived neurotrophic factor (BDNF);glial-derived neurotrophic factor (GDNF) and cAMP-responsive element-binding protein (CREB) between cells treated with fluoxetine for two weeks and their untreated counterparts. These results may help elucidate the mechanism of action of fluoxetine as a therapeutic drug for the treatment of depression. Data presented herein provide more evidence that, in addition to having a direct chemical effect on serotonin levels, fluoxetine can influence hESC-derived neuronal cells by increasing cell proliferation, while allowing them to maintain their neuronal characteristics.

무릎 반월연골판은 무릎에서 안정성과 충격 흡수능력, 그리고 축 성 압력을 받아주는 중요한 역할을 하고 있다. 각각의 반월연골 판은 환형 구조로 촘촘히 연결되어 있고, 듬성듬성 방사성 구조 로 결합되어 있으며, 무릎에 가해지는 축성 압력을 연골판 장력 (hoop tension)으로 변환시켜 준다. 실제로 체중의 약 50%-70%에 해당되는 하중이 반월연골판을 통해 전달되고 있다. 내측 반월연 골판 기시부 손상은 고리(hoop) 모양의 연속성이 끊어지게 되는 것으로 반월연골판의 이러한 생역학적 기능을 잃게 한다. 그리하 여 연골판 장력으로 변환되지 못한 축성 압력 때문에 무릎 관절 면의 퇴행성 변화를 초래한다고 알려져 있다. 이러한 반월연골판 병변의 진단 및 치료방침은 퇴행성 관절 염을 방지하는 관점에서 매우 중요하다. 내측 반월연골판 기시 부 파열은 크게 외상성 파열과 퇴행성 파열 두 가지로 나눌 수 있 다. 외상성 파열은 드물고 주로 연골판 기시부의 골부착부에서 떨어져 나오는 형태의 손상이며, 젊은 나이에서 전방십자인대 손 상과 잘 동반된다. 퇴행성 파열은 외상성 파열보다 비교적 흔하 며 기시부의 골부착부 자체가 아닌 골부착부 근처 실질에서의 파 열이 대부분이다. 50대 이상의 연령에서 여자에게 빈발하며, 큰 외상이 없이 퇴행성 변화로 생기는 파열이다. 한국에서의 통계 는 내측 반월연골판 파열의 약 25%를 차지할 정도로 수평파열 다 음으로 많다. 과거에는 동양에서 많이 발생한다고 알려졌으나 최근에는 서양에서도 많이 보고되고 있는데, 이는 관절경 검사와 자기공명영상(magnetic resonance imaging, MRI) 진단기술의 발달 pISSN : 1226-2102, eISSN : 2005-8918 372

Purpose: The anastomotic leakage rate after rectal resection has been reported to be approximately 2.5-21 percent, but most results were associated with open surgery. The aim of this study was to identify risk factors and their relationship to the experience of the surgeon for anastomotic leakage after laparoscopic rectal resection. Methods: Between March 2003 and December 2008, 156 patients underwent a laparoscopic rectal resection without a diverting ileostomy. The patients' characteristics, the details of treatment, the intraoperative results, and the postoperative results were recorded prospectively. Univariate and multivariate analyses were applied to identify risk factors for anastomotic leakage. Results: The majority of operations were performed for malignant disease (n = 150; 96.2%), and 96 patients (61.5%) were males. Conversion to open surgery occurred in 1 case (0.6%). The anastomotic leak rate was 10.3% (16/156), and there were no mortalities. In the univariate analysis, tumor location, anastomotic level, intraoperative events, and operation time were associated with increased anastomotic leakage rate. In the multivariate analysis, anastomotic level (odds ratio [OR], 6.855; 95% confidence interval [CI], 1.271 to 36.964) and operation time (OR, 8.115; 95% CI, 1.982 to 33.222) were significantly associated with anastomotic leakage. Conclusion: The important risk factors for anastomotic leakage after laparoscopic rectal resection without a diverting ileostomy were low anastomosis and long operation time. An additional procedure, such as diverting stoma, may reduce the anastomotic leakage if it is selectively applied in cases with these risk factors.