Integrative analysis of single-cell transcriptomics reveals age-associated immune landscape of glioblastoma.

Glioblastoma (GBM) is the most malignant tumor in center nervous system. Clinical statistics revealed that senior GBM patients had a worse overall survival (OS) comparing with that of patients in other ages, which is mainly related with tumor microenvironment including tumor-associated immune cells in particular. However, the immune heterogeneity and age-related prognosis in GBM are under studied. Here we developed a machine learning-based method to integrate public large-scale single-cell RNA sequencing (scRNA-seq) datasets to establish a comprehensive atlas of immune cells infiltrating in cross-age GBM. We found that the compositions of the immune cells are remarkably different across ages. Brain-resident microglia constitute the majority of glioblastoma-associated macrophages (GAMs) in patients, whereas dramatic elevation of extracranial monocyte-derived macrophages (MDMs) is observed in GAMs of senior patients, which contributes to the worse prognosis of aged patients. Further analysis suggests that the increased MDMs arisen from excessive recruitment and proliferation of peripheral monocytes not only lead to the T cell function inhibition in GBM, but also stimulate tumor cells proliferation via VEGFA secretion. In summary, our work provides new cues for the correlational relationship between the immune microenvironment of GBM and aging, which might be insightful for precise and effective therapeutic interventions for senior GBM patients.

Microglial Piezo1 senses Aß fibril stiffness to restrict Alzheimer's disease.

The pathology of Alzheimer's disease (AD) is featured with extracellular amyloid-ß (Aß) plaques, whose impact on the mechanical properties of the surrounding brain tissues is unclear. Microglia sense and integrate biochemical cues of the microenvironment. However, whether the microglial mechanosensing pathways influence AD pathogenesis is unknown. Here, we surveyed the elevated stiffness of Aß-plaque-associated tissues and observed the selective upregulation of the mechanosensitive ion channel Piezo1 in Aß-plaque-associated microglia. Piezo1 sensed the stiffness stimuli of Aß fibrils and subsequently induced Ca2+ influx for microglial clustering, phagocytosis, and compacting of Aß plaques. Microglia lacking Piezo1 led to the exacerbation of Aß pathology and cognitive decline, whereas pharmacological activation of microglial Piezo1 ameliorated brain Aß burden and cognitive impairment in 5 × FAD mice. Together, our results reveal that Piezo1, a mechanosensor of Aß fibril stiffness in microglia, represents a potential therapeutic target for AD.

Prognostic impact of tumor deposits on overall survival in colorectal cancer: Based on Surveillance, Epidemiology, and End Results database.

BACKGROUND: In colorectal cancer, tumor deposits (TDs) are considered to be a prognostic factor in the current staging system, and are only considered in the absence of lymph node metastases (LNMs). However, this definition and the subsequent prognostic value based on it is controversial, with various hypotheses. TDs may play an independent role when it comes to survival and addition of TDs to LNM count may predict the prognosis of patients more accurately. AIM: To assess the prognostic impact of TDs and evaluate the effect of their addition to the LNM count. METHODS: The patients are derived from the Surveillance, Epidemiology, and End Results database. A prognostic analysis regarding impact of TDs on overall survival (OS) was performed using Cox regression model, and other covariates associating with OS were adjusted. The effect of addition of TDs to LNM count on N restaging was also evaluated. The subgroup analysis was performed to explore the different profile of risk factors between patients with and without TDs. RESULTS: Overall, 103755 patients were enrolled with 14131 (13.6%) TD-positive and 89624 (86.4%) TD-negative tumors. TD-positive patients had worse prognosis compared with TD-negative patients, with 3-year OS rates of 47.3% (95%CI, 46.5%-48.1%) and 77.5% (95%CI, 77.2%-77.8%, P < 0.0001), respectively. On multivariable analysis, TDs were associated poorer OS (hazard ratio, 1.35; 95%CI, 1.31-1.38; P < 0.0001). Among TD-positive patients, the number of TDs had a linear negative effect on disease-free survival and OS. After reclassifying patients by adding TDs to the LNM count, 885 of 19 965 (4.4%) N1 patients were restaged as pN2, with worse outcomes than patients restaged as pN1 (3-year OS rate: 78.5%, 95%CI, 77.9%-79.1% vs 63.2%, 95%CI, 60.1%-66.5%, respectively; P < 0.0001). CONCLUSION: TDs are an independent prognostic factor for OS in colorectal cancer. The addition of TDs to LNM count improved the prognostic accuracy of tumor, node and metastasis staging.

Predictive value of serum alpha-fetoprotein for tumor regression after preoperative chemotherapy for rectal cancer.

BACKGROUND: Preoperative therapy is widely used in locally advanced rectal cancer. It can improve local control of rectal cancer. However, there are few indicators that can predict the effect of preoperative chemotherapy accurately. AIM: To investigate whether the increase in serum α-fetoprotein (AFP) can predict better efficacy of preoperative chemotherapy. METHODS: This was a retrospective study. We analyzed 125 patients admitted between 2017 and 2019 with locally advanced rectal cancer. All patients received six cycles of preoperative chemotherapy (mFOLFOX6 every 2 wk). Serum AFP of 26 patients rose slightly after three or four cycles of chemotherapy, and fell to normal again within 2 mo. The other 99 patients had a normal level of serum AFP during chemotherapy. Patients were divided into two groups (AFP risen and AFP normal). According to postoperative pathology, we compared tumor regression and complete response rate between the two groups. The primary outcome measure was the tumor regression grade (TRG) after chemotherapy. The difference in pathological complete response between the two groups was also investigated. RESULTS: There were no tumor progression and distant metastasis in both groups during preoperative chemotherapy. Patients in the AFP risen group achieved better TRG 0/1 than those in the AFP normal group (61.5% vs 39.4%). The increase in AFP was a significant predictor for better tumor regression [χ 2 = 4.144, odds ratio (OR) = 2.666, P = 0.04]. In the AFP risen group, the complete response rate was 30.8%, which was higher than in the AFP normal group (30.8% vs 12.1%, χ 2 = 4.542, OR = 3.251, P = 0.03). CONCLUSION: Patients with a slight increase in serum AFP can achieve better tumor regression during preoperative chemotherapy, and are more likely to achieve pathological complete response.

The Oligodendrocyte Transcription Factor 2 OLIG2 regulates transcriptional repression during myelinogenesis in rodents.

OLIG2 is a transcription factor that activates the expression of myelin-associated genes in the oligodendrocyte-lineage cells. However, the mechanisms of myelin gene inactivation are unclear. Here, we uncover a non-canonical function of OLIG2 in transcriptional repression to modulate myelinogenesis by functionally interacting with tri-methyltransferase SETDB1. Immunoprecipitation and chromatin-immunoprecipitation assays show that OLIG2 recruits SETDB1 for H3K9me3 modification on the Sox11 gene, which leads to the inhibition of Sox11 expression during the differentiation of oligodendrocytes progenitor cells (OPCs) into immature oligodendrocytes (iOLs). Tissue-specific depletion of Setdb1 in mice results in the hypomyelination during development and remyelination defects in the injured rodents. Knockdown of Sox11 by siRNA in rat primary OPCs or depletion of Sox11 in the oligodendrocyte lineage in mice could rescue the hypomyelination phenotype caused by the loss of OLIG2. In summary, our work demonstrates that the OLIG2-SETDB1 complex can mediate transcriptional repression in OPCs, affecting myelination.

Temporarily Epigenetic Repression in Bergmann Glia Regulates the Migration of Granule Cells.

Forming tight interaction with both Purkinje and granule cells (GCs), Bergmann glia (BG) are essential for cerebellar morphogenesis and neuronal homeostasis. However, how BG act in this process is unclear without comprehensive transcriptome landscape of BG. Here, high temporal-resolution investigation of transcriptomes with FACS-sorted BG revealed the dynamic expression of genes within given functions and pathways enabled BG to assist neural migration and construct neuron-glia network. It is found that the peak time of GCs migration (P7-10) strikingly coincides with the downregulation of extracellular matrix (ECM) related genes, and the disruption of which by Setdb1 ablation at P7-10 in BG leads to significant migration defect of GCs emphasizing the criticality of Nfix-Setdb1 mediated H3K9me3 repressive complex for the precise regulation of GCs migration in vivo. Thus, BG's transcriptomic landscapes offer an insight into the mechanism by which BG are in depth integrated in cerebellar neural network.

Decoupling and detecting angular motion errors based on a line laser sensor for motion platforms.

Efficient measurement of a line laser sensor can be achieved by combining it with a multi-axis platform. However, the measuring results are significantly affected by the precision of the platform. To provide a criterion for verifying and compensating for the geometric errors of a platform, a measuring method based on a line laser sensor for angular motion errors is proposed for the first time to the best of our knowledge in this work. The principles of separating and simultaneously detecting the yaw, roll, and pitch errors are elaborated. Based on this, the detection model is built for a three-dimensional platform. Under such an experimental condition, the stability and accuracy of the line laser sensor is tested, and the experimental results revealed small deviations of 3 µm and 10 µm in z axis and x axis, respectively. Ultimately, a series of experiments is performed to verify the feasibility and repeatability of the method. The yaw experimental results indicate that the trend and rule of error conform to the measured value by a laser interferometer, and the maximum variation value is 0.0075 mrad.

Viscoelastic characterization of injured brain tissue after controlled cortical impact (CCI) using a mouse model.

BACKGROUND: Mechanical properties of the brain tissue are crucial to understand the mechanisms of traumatic brain injury (TBI). Injured brain tissue could induce changes of mechanical properties and anatomical structures. However, limited data is available for the injured tissue. NEW METHOD: We developed a custom-built device to introduce controlled cortical impact (CCI) to brain with controlled impact velocity and direction. A study protocol for measuring the viscoelastic properties of injured brain tissue was also developed. Micro-scale morphological changes of the vasculature were quantified by analyzing confocal images of the brain tissue using CLARITY method. RESULTS: Results showed significant differences of the instantaneous shear modulus of the impact region from different impact angles. However, no significant differences were found for long-term shear modulus by varying the impact angles and velocities. Analysis of the vasculature showed an increased radius of the vessels in the injured tissue compared with that in the control group. COMPARISON WITH EXISTING METHODS: A combination of three different impact velocities and three different impact angles were adopted for producing injury to the brain. In addition, viscoelastic properties were compared between the injured and non-injured regions. The corresponding morphological changes of the vasculature system were also investigated. CONCLUSIONS: The instantaneous shear modulus at the impact region was significantly different for the three impact angles. Compared to that of the control group, increased radius of the vasculature was also observed in the injured brain tissue. Results indicated that the biomechanical and structural changes of the injured tissue were closely related to the impact angles and velocities. Viscoelastic measurements could also help validation of computational models.

Nucleoporin Seh1 Interacts with Olig2/Brd7 to Promote Oligodendrocyte Differentiation and Myelination.

Nucleoporins (Nups) are involved in neural development, and alterations in Nup genes are linked to human neurological diseases. However, physiological functions of specific Nups and the underlying mechanisms involved in these processes remain elusive. Here, we show that tissue-specific depletion of the nucleoporin Seh1 causes dramatic myelination defects in the CNS. Although proliferation is not altered in Seh1-deficient oligodendrocyte progenitor cells (OPCs), they fail to differentiate into mature oligodendrocytes, which impairs myelin production and remyelination after demyelinating injury. Genome-wide analyses show that Seh1 regulates a core myelinogenic regulatory network and establishes an accessible chromatin landscape. Mechanistically, Seh1 regulates OPCs differentiation by assembling Olig2 and Brd7 into a transcription complex at nuclear periphery. Together, our results reveal that Seh1 is required for oligodendrocyte differentiation and myelination by promoting assembly of an Olig2-dependent transcription complex and define a nucleoporin as a key player in the CNS.

Deficits in Bone Geometry in Growth Hormone-Deficient Prepubertal Boys Revealed by High-Resolution Peripheral Quantitative Computed Tomography.

INTRODUCTION: Although growth hormone (GH) is essential for attainment of peak bone mass, bone health in prepubertal children with GH deficiency is not routinely evaluated. The objective of this study was to evaluate bone microarchitecture in GH-deficient (GHD) boys using high-resolution peripheral quantitative computed tomography (HR-pQCT). METHODS: Fifteen control and fifteen GHD, GH naïve pre-pubertal boys were recruited for a case-control study at a major academic center. Subjects with panhypopituitarism, chromosomal pathology, chronic steroids, or stimulant use were excluded. Volumetric bone mineral density (vBMD; total, cortical, and trabecular), bone geometry (total, cortical and trabecular cross-sectional area, cortical perimeter), bone microarchitecture, and estimated bone strength of the distal radius and tibia were assessed by HR-pQCT. Areal BMD and body composition were assessed by DXA. Insulin-like growth factor 1 (IGF-1), osteocalcin, C telopeptide, and P1NP levels were measured. RESULTS: GHD subjects had a significantly smaller cortical perimeter of the distal radius compared to controls (p < 0.001), with the difference in cortical perimeter persisting after adjusting for height z score, age, lean mass, and 25-hydroxyvitamin D level (p < 0.05).No significant differences were found in vBMD. No significant differences were found in microarchitecture, estimated strength, areal BMD, body composition, or bone turnover markers. Analysis showed significant positive correlations between IGF-1 levels and cortical parameters. DISCUSSION/CONCLUSIONS: Prepubertal GHD boys had deficits in bone geometry not evident with DXA. Larger prospective/longitudinal HR-pQCT studies are needed to determine the extent of these deficits, the need for routine bone evaluation, and the timing of GH replacement for prevention or restoration of these deficits.

ULK1/2 Constitute a Bifurcate Node Controlling Glucose Metabolic Fluxes in Addition to Autophagy.

Metabolic reprogramming is fundamental to biological homeostasis, enabling cells to adjust metabolic routes after sensing altered availability of fuels and growth factors. ULK1 and ULK2 represent key integrators that relay metabolic stress signals to the autophagy machinery. Here, we demonstrate that, during deprivation of amino acid and growth factors, ULK1/2 directly phosphorylate key glycolytic enzymes including hexokinase (HK), phosphofructokinase 1 (PFK1), enolase 1 (ENO1), and the gluconeogenic enzyme fructose-1,6-bisphosphatase (FBP1). Phosphorylation of these enzymes leads to enhanced HK activity to sustain glucose uptake but reduced activity of FBP1 to block the gluconeogenic route and reduced activity of PFK1 and ENO1 to moderate drop of glucose-6-phosphate and to repartition more carbon flux to pentose phosphate pathway (PPP), maintaining cellular energy and redox homeostasis at cellular and organismal levels. These results identify ULK1/2 as a bifurcate-signaling node that sustains glucose metabolic fluxes besides initiation of autophagy in response to nutritional deprivation.

Effects of colonic electrical stimulation using different individual parameter patterns and stimulation sites on gastrointestinal transit time, defecation, and food intake.

PURPOSE: This study aimed to compare the effects of colonic electrical stimulation (CES) on gastrointestinal transit time (GITT), energy consumption, stool frequency, stool consistency, and food intake using different individual parameter patterns and stimulation sites. METHODS: Eight beagle dogs underwent surgery and CES. First, CES was conducted to determine the individual parameters with different pulse configurations, based on symptoms. Second, influences on energy consumption and GITT were compared between CES sessions with different pulse configurations. Third, GITT, stool frequency, stool consistency, and food intake were compared to assess the effects of CES at different stimulation sites. RESULTS: The individual parameters varied greatly among the dogs. In proximal colon electrical stimulation (PCES) and rectosigmoid colon electrical stimulation (RCES), energy consumption was lower with the constant pulse width mode than with the constant pulse amplitude mode (p = 0.012 and p = 0.018, respectively). There was no statistical difference between the two pulse configurations in GITT assessment. The PCES, RCES, and sequential CES sessions significantly accelerated GITT compared to sham stimulation. There was no statistical difference in GITT between PCES, RCES, and sequential CES sessions. Compared to sham CES session, RCES and sequential CES sessions exhibited significant higher stool frequency (p < 0.001 and p = 0.001, respectively), and PCES and RCES sessions inhibited food intake (p = 0.003 and p = 0.002, respectively). CONCLUSIONS: Constant pulse width mode is an appropriate pulse configuration for individual CES. At different stimulation sites, CES may exert different effects on stool frequency and food intake. This study provides an experimental basis for the clinical application of CES.

Retinoic acid morpholine amide (RAMA) inhibits expression of Fas ligand through EP1 receptor in colon cancer cells.

Among the members of tumour necrosis factor family Fas ligand on binding to its receptor strongly induces apoptosis of tumour-infiltrating lymphocytes (TIL). Thus, FasL acts as an inhibitor of anti-tumour immune response. The present study demonstrates that retinoic acid morpholine amide (RAMA) significantly suppresses FasL expression in colon cancer cells in a dose- and time-dependent manner. The suppression of FasL mRNA and proteins was significant at a concentration of 30 µM after 48 h in CLT85 and HT26 colon cancer cells. There was around 2.6- and 3.2-fold decrease in FasL mRNA after incubation with 30 µM of RAMA in CLT85 cells and HT26 cells, respectively. The results from Western blot showed a decrease in FasL mRNA and protein expression in both CLT85 and HT26 cells after suppression of cyclooxygenase (COX)-2 and COX-1 by RNAi. However, when COX-2-specific silencer RNA (siCOX-2)- and siCOX-1-treated CLT85 and HT26 cells were exposed to RAMA, inhibition of FasL expression was further suppressed. The siCOX-2-treated CLT85 and HT26 cells on exposure to RAMA showed ∼87 and ∼54 % reduction in FasL mRNA, respectively. Co-culture of Jurkat T cells with RAMA-treated HT26 and CLT85 cells decreased the viability of Jurkat T cells by only 2 and 4.3 %, respectively, compared to 19.5 and 37.3 % in control HT26 and CLT85 cells. The results from real-time reverse transcription polymerase chain reaction (RT-PCR) and immunoblotting showed that suppression of EP1 prevented RAMA-induced FasL suppression in CLT85 cells at both the mRNA and protein levels. Thus, RAMA can be a potent therapeutic agent for the treatment of colon tumours.

The effects of individualized gastric electrical stimulation on food craving and gastrointestinal peptides in dogs.

BACKGROUND: Using an adjustable stimulator with a wide range of stimulation parameters, the aims of this study were 1) to investigate the effects of long-term gastric electrical stimulation (GES) on appetite and differential food cravings for three different foods and 2) to investigate the effects of GES on plasma gastrointestinal peptide concentrations. METHODS: The study was performed in eight Beagle dogs implanted with one pair of serosal electrodes. They were followed during GES and sham GES sessions in a crossover design. GES was conducted using a series of individualized parameters. Food intake and food cravings were observed to evaluate the effects of long-term GES. Enzyme-linked immunosorbent assay was used to measure the plasma concentrations of gastrointestinal peptides. RESULTS: Dogs on GES for three months ate significantly less food than those on sham GES for three months (p < 0.05). A significant change in food cravings was induced by GES. Dogs with GES ate significantly less high-fat food. However, there was no significant difference in consumption of high-carbohydrate food or balanced food between the periods of GES and sham GES. The plasma concentrations of ghrelin, peptide YY3-36, and glucagon-like peptide 1 did not differ significantly between the periods of GES and sham GES. CONCLUSIONS: Food intake and food craving were changed significantly by adjustable GES. GES may be used for treating obesity by changing food preferences. Further clinical studies are necessary to highlight the effect of adjustable GES on eating behavior.

The acute effects of a new type of implantable gastric electrical stimulators featuring varied pulse widths on beagle dogs' food intake and gastric accommodation.

BACKGROUND: To improve the therapeutic effects of gastric electrical stimulation (GES) for obesity, an animal experiment was conducted using a new type of stimulators. Proper parameters of GES were selected, and the impacts of GES on the food intake and gastric accommodation of canines were observed. METHODS: Eight beagle dogs were operated on, and GES was performed on them. Firstly, GES was performed to determine the right parameters according to symptoms. Secondly, the so selected parameters were used in a 3-day GES procedure, during which process food intake, body weight, and symptoms were recorded. Thirdly, the gastric capacities before and after GES with different pulse widths were measured by means of a barostat. RESULTS: The selected parameters varied for each dog, with the pulse widths ranging from 0.3 to 6 ms. The food consumption after GES dropped significantly as compared with the amount observed in the sham stimulation. Tolerance to stimulation could be observed during GES. The post-GES gastric fundus capacity increased evidently in comparison with the capacity before GES, suggesting significant distention as compared with sham stimulation. Given an increment of 2 ms in the pulse width twice, the gastric capacity continued to distend each time. CONCLUSIONS: GES featuring pulse trains with wider and individualized pulse widths could inhibit food consumption of dogs. The stimulation parameters should be selected individually and adjusted periodically. GES of this mode could also increase the fasting gastric capacity with certain dose-related effects. The new type of stimulators may be more suitable for the treatment of human obesity than traditional stimulators.

Parameter selection and stimulating effects of an adjustable gastric electrical stimulator in dogs.

BACKGROUND: Gastric electrical stimulation (GES) has been proposed as a promising therapeutic option in treating obesity for 20 years. Currently, the available device of GES cannot meet the clinical needs. The purpose of this study is to verify the effect of a new type of adjustable gastric electrical stimulator in reducing food intake and body weight. METHODS: Eight beagle dogs randomly followed GES and sham GES for 3 months in a crossover design. Parameters were adjusted and individualized during the experiment. Symptoms of GES were recorded, and the effective parameters were selected. Resistance to GES was assessed. Food intake and body weight were measured to evaluate the effect of GES. RESULTS: The effective parameters were varied among the dogs. Resistance to GES was observed in different periods in dogs. Parameters needed to be adjusted every 10.2 ± 2.1 days during the period of GES. Food intake during GES for 3 months was significantly reduced than that during sham GES of 3 months (P < 0.05). With the decreased food intake, body weight was significantly reduced by the end of GES of 3 months compared with that of sham GES of 3 months (P < 0.05). CONCLUSIONS: Food intake and body weight of dogs are significantly reduced by adjustable GES. Individual parameters and resistance during GES are required to be considered. The new adjustable device may have good prospects of clinical application for obesity.

[Preoperative evaluation of mesenteric vascular anatomy using 256 multi-slice computed tomography before laparoscopic surgery].

OBJECTIVE: To evaluate mesenteric vascular anatomy using 256 multi-slice computed tomography (MSCT) before laparoscopic colorectal surgery. METHODS: Eleven patients with colorectal cancer underwent 256 MSCT from February 2010 to December 2010. The evaluation items were visualization of mesenteric artery and vein by 3-dimensional CT angiography, which was compared with findings on laparoscopic surgery. RESULTS: Three-dimensional CT angiography correctly demonstrated variations of the mesenteric artery and vein and were consistent with laparoscopic findings. Of the 3 patients undergoing right hemicolectomy, ileocolic artery (ICA) ran ventrally to the superior mesenteric vein(SMV) in 1 patient, whereas ICA ran dorsally to the SMV in 2 patients; the right colic artery (RCA) branched directly from superior mesenteric artery(SMA) in 2 patients; RCA was absent and the left branch of middle colic artery(MCA) fed the tumor in 1 patient. In the patients who had transverse colon resection, MCA branched from SMA. In 2 of 3 patients who underwent sigmoidectomy, sigmoid artery (SA) branched from left colic artery(LCA); in 1 of 3 patients of sigmoid resection, SA branched from inferior mesenteric artery(IMA). In 4 patients with rectal cancer, the superior rectal artery (SRA) fed the tumor. CONCLUSION: The 256 MSCT is effective for evaluating mesenteric vascular anatomy variation before laparoscopic surgery for colorectal cancer.

[Experimental study on the stimulating effect and the tissue compatibility of a new type of implanted gastric electrical stimulator].

A new type of gastric electrical stimulator (GES) was introduced. After the stimulator was implanted in beagle dogs, its stimulating effects and the pathological changes at the implant site were observed to study the safety and efficacy of stimulator as well as the tissue compatibility of the materials used. The results showed that, this type of stimulator was safe and capable of inhibiting food intake of the dogs, and that the materials used had good tissue compatibility.