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Introduction Transcription factors (TFs) and cis-regulatory elements (CREs) control gene transcripts involved in various biological processes. We hypothesize that TFs and CREs can be effective molecular tools for De Novo regulation designs to engineer plants. Objectives We selected two Arabidopsis TF types and two tobacco CRE types to design a De Novo regulation and evaluated its effectiveness in plant engineering. Methods G-box and MYB recognition elements (MREs) were identified in four Nicotiana tabacum JAZs (NtJAZs) promoters. MRE-like and G-box like elements were identified in one nicotine pathway gene promoter. TF screening led to select Arabidopsis Production of Anthocyanin Pigment 1 (PAP1/MYB) and Transparent Testa 8 (TT8/bHLH). Two NtJAZ and two nicotine pathway gene promoters were cloned from commercial Narrow Leaf Madole (NL) and KY171 (KY) tobacco cultivars. Electrophoretic mobility shift assay (EMSA), cross-linked chromatin immunoprecipitation (ChIP), and dual-luciferase assays were performed to test the promoter binding and activation by PAP1 (P), TT8 (T), PAP1/TT8 together, and the PAP1/TT8/Transparent Testa Glabra 1 (TTG1) complex. A DNA cassette was designed and then synthesized for stacking and expressing PAP1 and TT8 together. Three years of field trials were performed by following industrial and GMO protocols. Gene expression and metabolic profiling were completed to characterize plant secondary metabolism. Results PAP1, TT8, PAP1/TT8, and the PAP1/TT8/TTG1 complex bound to and activated NtJAZ promoters but did not bind to nicotine pathway gene promoters. The engineered red P + T plants significantly upregulated four NtJAZs but downregulated the tobacco alkaloid biosynthesis. Field trials showed significant reduction of five tobacco alkaloids and four carcinogenic tobacco specific nitrosamines in most or all cured leaves of engineered P + T and PAP1 genotypes. Conclusion G-boxes, MREs, and two TF types are appropriate molecular tools for a De Novo regulation design to create a novel distant-pathway cross regulation for altering plant secondary metabolism.

Background and aim Westernized high-fat diet increases the risk for inflammatory bowel diseases (IBDs), yet with insufficient understanding of the role of high-protein diet. We aimed to identify the effect of high-protein diets from different dietary proteins (casein, whey protein, soy protein) on experimental colitis and its impact on microbiota, structure and function of colonic mucus layer. Methods Female BALB/c mice were fed by standard diet, high-casein diet (HCD), high whey protein diet or high soy protein diet for 4 weeks. The susceptibility of dextran sulfate sodium (DSS)-induced colitis in mice and thickness of colonic mucus layer were compared after different dietary interventions, associated with the identification of the reversal effect of broad-spectrum antibiotic intervention (0.5 g/L of vancomycin and 1 g/L of neomycin sulfate, metronidazole and ampicillin in drinking water). Further analysis was performed on the synthesis of mucin, microbiota and sialidase involved in degradation of mucus layer. Results High-protein diets aggravated acute DSS-induced colitis independent of protein composition, while broad-spectrum antibiotics reversed this effect. HCD significantly altered the composition of bacteria in the colonic mucus layer, especially Bacteroides thetaiotaomicron and total mucin-degrading bacteria; besides, it increased sialidase concentration and reduced the thickness of mucus layer. However, it exhibited no significant effect on the synthesis of Muc2. Broad-spectrum antibiotics decreased the abundance of mucin-degrading bacteria and sialidase concentration while increased the thickness of mucus layer. Conclusion High-protein diet shifts microbial composition and thickness of colonic mucus layer, leading to the aggravation of acute DSS-induced colitis.

It is difficult to control the balance between artifact suppression and detail preservation. In addition, the information contained in the reconstructed image is limited. For achieving the purpose of less lost information and lower computational complexity in the sampling process, this paper proposed a novel algorithm to realize the image reconstruction using sparse representation. Firstly, the principle of algorithm for sparse representation is introduced, and then the current commonly used reconstruction algorithms are described in detail. Finally, the algorithm can still process the image when the sparsity is unknown by introducing the sparsity theory and dynamically changing the step size to approximate the sparsity. The results explain that the improved algorithm can not only reconstruct the image with unknown sparsity, but also has advantages over other algorithms in reconstruction time. In addition, compared with other algorithms, the reconstruction time of the improved algorithm is the shortest under the same sampling rate.

The dysfunction of placenta development is correlated to the defects of pregnancy and fetal growth. The detailed molecular mechanism of placenta development is not identified in humans due to the lack of material in vivo. Trophoblast (TB) lineage derived from human embryonic stem cells (hESCs) induced by bone morphogenetic protein 4 (BMP4) has been applied as a model for studying TB lineage specification in vitro. With the development of single-cell sequencing technology, it became possible to detect the transcriptome of the post-implantation embryo at unprecedented precision. In this study, we reanalyzed single-cell RNA-seq of post-implantation embryos derived from two separate groups and identified different subtypes of trophoblast cells and their marker, respectively. At the same time, we focused on the gene expression patterns of trophoblast-specific transcription factors in different models. Our analysis sheds new light on the transcription regulation mechanism of trophoblast differentiation at the early stage of pregnancy establishment in human.

The use of models of stem cell differentiation to trophoblastic cells provides an effective perspective for understanding the early molecular events in the establishment and maintenance of human pregnancy. In combination with the newly developed deep learning technology, the automated identification of this process can greatly accelerate the contribution to relevant knowledge. Based on the transfer learning technique, we used a convolutional neural network to distinguish the microscopic images of Embryonic stem cells (ESCs) from differentiated trophoblast -like cells (TBL). To tackle the problem of insufficient training data, the strategies of data augmentation were used. The results showed that the convolutional neural network could successfully recognize trophoblast cells and stem cells automatically, but could not distinguish TBL from the immortalized trophoblast cell lines in vitro (JEG-3 and HTR8-SVneo). We compare the recognition effect of the commonly used convolutional neural network, including DenseNet, VGG16, VGG19, InceptionV3, and Xception. This study extends the deep learning technique to trophoblast cell phenotype classification and paves the way for automatic bright-field microscopic image analysis of trophoblast cells in the future.

Background Relapse is a major obstacle in the treatment of acute myeloid leukemia (AML). Refinement of risk stratification may aid the identification of patients who are likely to relapse. Abnormal cysteine and glycine-rich protein 2 (CSRP2) has been implicated in various cancers, but its function remains unclear. The purpose of this study was to explore the role of CSRP2 in predicting adult AML recurrence. Methods RT-PCR was used to detect the expression of CSRP2 in 193 newly diagnosed adult AML patients and 44 healthy controls. The competitive risk model was used to calculate the cumulative incidence of relapse rate (CIR), Kaplan–Meier to calculate the relapse-free survival rate (RFS), and the Cox regression model to perform multivariate analysis. Viral transfection was used to construct AML cell lines with stable knockdown of CSRP2, CCK8 to detect proliferation and drug resistance, flow cytometry to detect cell cycle and apoptosis, and Western blot to detect key molecules in signaling pathways. Results CSRP2 transcript levels were higher in 193 adult AML compared with 44 healthy controls. In 149 patients who achieved complete remission, those with high CSRP2 transcript levels displayed a lower 2-year CIR and higher 2-year RFS, especially when receiving only chemotherapy. In multivariate analysis, a high CSRP2 transcript level was independently associated with a better RFS. Knockdown of CSRP2 promoted proliferation and cell cycle progression, and reduced chemosensitivity. Western blot analysis showed upregulation of p-AKT and p-CREB in CSRP2-knockdown AML cell lines. Inhibition assays suggested these two signaling pathways participated in the CSRP2-mediated proliferation effects in AML cell lines. Conclusion In summary, CSRP2 correlates with relapse in adult AML. Down-regulation of CSRP2 could promote the proliferation of AML cell lines by regulating the AKT and CREB signaling pathways. Therefore, CSRP2 may provide prognostic significance and potential therapeutic targets in the management of AML.

Background Glioma is a common brain malignancy with high mortality. The competing endogenous RNA (ceRNA) networks may play key roles in cancer progression. This study was conducted to probe the role of long noncoding RNA (lncRNA) NCK1-AS1 in glioma progression and the involved mechanisms. Methods Microarray analyses were performed to explore the lncRNAs/miRNAs/genes with differential expression in glioma. NCK1-AS1 levels in glioma tissues and normal brain tissues, and in glioma cell lines and normal human glial cells were identified. The interactions among NCK1-AS1, miR-138-2-3p and TRIM24 were validated through luciferase reporter, RNA immunoprecipitation and RNA pull-down assays. Gain- and loss-of functions of NCK1-AS1, miR-138-2-3p and TRIM24 were performed to identify their roles in the behaviors of glioma cells. The activity of the Wnt/β-catenin pathway was measured. In vivo experiments were performed as well. Results High expression of NCK1-AS1 was found in glioma tissues and cells, especially in U251 cells. Online predictions and the integrated experiments identified that NCK1-AS1 elevated the TRIM24 expression through sponging miR-138-2-3p, and further activated the Wnt/β-catenin pathway. Artificial silencing of NCK1-AS1 or up-regulation of miR-138-2-3p led to inhibited proliferation, invasion and migration but promoted cell apoptosis of U251 cells, while up-regulation of TRIM24 reversed these changes, and it activated the Wnt/β-catenin pathway. The in vitro results were reproduced in in vivo experiments. Conclusions Our study suggested that NCK1-AS1 might elevate TRIM24 expression and further activate the Wnt/β-catenin pathway via acting as a ceRNA for miR-138-2-3p. Silencing of NCK1-AS1 might inhibit the progression of glioma.

STUDY DESIGN Prospective, randomized, controlled trial. OBJECTIVE To compare robot-assisted and conventional implantation techniques by evaluating the accuracy and safety of implanting screws in cervical vertebrae. SUMMARY OF BACKGROUND DATA Cervical spinal surgery is difficult and dangerous as screw misplacement might lead not only to decreased stability but also neurological, vascular, and visceral injuries. A new robot-assisted surgical procedure has been introduced to improve the accuracy of implant screw positioning. METHODS We randomly assigned 135 patients with newly diagnosed cervical spinal disease and who required screw fixation using either robot-assisted or conventional fluoroscopy-assisted cervical spinal surgery. The primary outcomes were the discrepancies between the planned trajectories and the actual screw positions. RESULTS Altogether, 127 patients underwent the assigned intervention (61 robot-assisted and 66 conventional fluoroscopy-assisted). The baseline characteristics including the screw types, were similar in the two groups. Altogether, 390 screws were planed and placed in the cervical vertebrae, and 94.9% were acceptable. The robot-assisted group had a better screw placement accuracy than the conventional fluoroscopy-assisted group with associated P values

Spinal cord injury (SCI) is the major cause of the spinal damage affecting motor and sensory function. Thus, the present study was conducted to investigate the effect of D-pinitol (PN) on spinal cord injury in rats. The PN showed to recover motor function near to normal via modulating oxidative stress, inflammatory response and cellular apoptosis in SCI rats. PN also causes modulation of Bcl2 family proteins and reduces the level of NF-ĸB and LOX-1 in dose dependent manner. The PN causes reduction of NLRP3, TNF-α and iNOS, with increase in caspase-1 together with modulation of MAPK mediators. It has been suggested that, D-pinitol exert protective action against SCI via inhibition of apoptosis, inflammation and oxidative stress, via modulating Bcl-2 genes and MAPK pathway.

G-quadruplexes are the non-canonical nucleic acid structures that are preferentially formed in G-rich regions. This structure has been shown to be associated with many biological functions. Regardless of the broad efforts on DNA G-quadruplexes, we still have limited knowledge on RNA G-quadruplexes, especially in a transcriptome-wide manner. Herein, by integrating the DMS-seq and the bioinformatics pipeline, we profiled and depicted the RNA G-quadruplexes in the human transcriptome. The genes that contain RNA G-quadruplexes in their specific regions are significantly related to immune pathways and the COVID-19-related gene sets. Bioinformatics analysis reveals the potential regulatory functions of G-quadruplexes on miRNA targeting at the scale of the whole transcriptome. In addition, the G-quadruplexes are depleted in the putative, not the real, PAS-strong poly(A) sites, which may weaken the possibility of such sites being the real cleaved sites. In brief, our study provides insight into the potential function of RNA G-quadruplexes in post-transcription.

Qing-Chang-Hua-Shi (QCHS) is a Chinese herbal formula, which is composed of 11 herbs. Studies have also shown that QCHS granules can alleviate colitis in animal models by preventing inflammatory responses and suppressing apoptosis through the MEK/ERK signaling pathway. To determine the efficacy and safety of QCHS granules in patients with moderately active UC. We performed a multicenter, randomized, placebo-controlled, double-blind study of patients with moderately active UC who did not respond to 4 weeks of mesalazine therapy at the maximum dose. Patients were randomly assigned to groups and administered QCHS granules (125 g/day, n = 59) or an identical placebo, which was similar to the QCHS granules in color and taste (125 g/day, n = 60), with continued 5-ASA 4 g/d therapy for 12 weeks. The primary outcome was the rate of clinical response and clinical remission at week 12. The secondary outcomes were health-related quality of life, endoscopic response rate, and mucosal healing rate. Any changes in mucus/bloody stool and diarrhea were recorded. Out of the 119 enrolled patients at 10 different centers in China, 102 patients completed the trial. Clinical remission and clinical response were seen in 31.48% and 92.59% of QCHS-treated patients, and 12.50% and 72.92% of placebo-treated patients, respectively. There was a significant difference between the two treatment groups. More patients receiving QCHS granules vs. placebo achieved remission of mucus/bloody stool (70.37% vs. 47.92%, P = 0.0361). Adverse event rates were similar (QCHS granules 38.33%; placebo 25.42%). In conclusion, QCHS granules were superior to the placebo in introducing clinical remission and mucosal healing, as well as in relieving mucus/blood stool in patients with moderately active and 5-ASA-refractory UC.

Immunosuppression is critical for tumor growth and metastasis as well as obstacle to effective immunotherapy. Here, we demonstrate that host deficiency in caveolin-2, a member of caveolin protein family, increases M1-polarized tumor-associated macrophage (TAM) and CD8 T cell infiltration into subcutaneously implanted murine lung carcinoma tumors. Importantly, increase in M1 TAM-specific markers and cytokines occurs prior to increased numbers of tumor-infiltrating CD8 T cells and tumor regression in caveolin-2 deficient mice, suggesting that an early increase in M1 TAMs is a novel mechanism, via which host deficiency in caveolin-2 inhibits tumor growth. Consistent with the latter, transfer and co-injection of caveolin-2 deficient bone marrow (origin of TAMs) suppresses tumor growth and increases numbers of M1-polarized TAMs in wild type mice. Collectively, our data suggest that lung cancer cells use caveolin-2 expressed in bone marrow-derived cell types including TAMs to promote tumor growth via suppressing the anti-tumor immune response and that caveolin-2 could be a potential target for cancer immunotherapy.

The plant flavonoid dogma proposes that labile plant flavonoid carbocations (PFCs) play vital roles in the biosynthesis of proanthocyanidins (PAs). However, whether PFCs exist in plants and how PFCs function remain unclear. Here, we report the use of an integrative strategy including enzymatic assays, mutant analysis, metabolic engineering, isotope labeling and metabolic profiling to capture PFCs and demonstrate their functions. In anthocyanidin reductase (ANR) assays, an (-)-epicatechin conjugate was captured in protic polar nucleophilic methanol alone or methanol-HCl extracts. Tandem mass spectrum (MS/MS) analysis characterized this compound as an (-)-epicatechin-4-O-methyl (EOM) ether, which resulted from (-)-epicatechin carbocation and the methyl group of methanol. Acid-based catalysis of procyanidin B2 and B3 produced four compounds, which were annotated as two EOM and two (+)-catechin-4-O-methyl (COM) ethers. Metabolic profiling of seven PA pathway mutants showed an absence or reduction of two EOM ether isomers in seeds. Camellia sinensis ANRa (CsANRa), leucoanthocyanidin reductase c (CsLARc), and CsMYB5b (a transcription factor) were independently overexpressed for successful PA engineering in tobacco. The EOM ether was remarkably increased in CsANRa and CsMYB5b transgenic flowers. Further metabolic profiling for eight green tea tissues revealed two EOM and two COM ethers associated with PA biosynthesis. Moreover, an incubation of (-)-epicatechin or (+)-catechin with epicatechin carbocation in CsANRa transgenic flower extracts formed dimeric procyanidin B1 or B2, demonstrating the role of flavan-3-ol carbocation in the formation of PAs. Taken together, these findings indicated that flavan-3-ol carbocations exist in extracts and are involved in the biosynthesis of PAs of plants.

There is growing evidence of the position of microRNAs (miRs) in Alzheimer's disease (AD), thus our objective was to discuss the impact of miR-129-5p regulating nerve injury and inflammatory response in AD rats by modulating SOX6 expression. The AD rat model was established by injecting Aβ25-35 into the brain. The pathological changes, ultrastructure, number of neurons, cell degeneration and apoptosis of hippocampal tissue were observed in vivo. MiR-129-5p, SOX6, IL-1β, TNF-α, Bcl-2 and Bax expression in serum and hippocampal tissues were detected by ELISA, RT-qPCR or western blot analysis. The successfully modeled hippocampal neuronal cells of AD were transfected with miR-129-5p mimic, SOX6-siRNA or their controls to figure out their roles in proliferation, apoptosis and inflammatory reaction in vitro. Low expression of SOX6 and high expression of miR-129-5p in vivo of rats would shorten the escape latent period and increase the times of crossing platforms, alleviate the pathological injury, inhibit neuronal apoptosis and reduce the inflammatory reaction. Up-regulation of miR-129-5p and down-regulation of SOX6 promoted proliferation, suppressed apoptosis and degraded the inflammatory reaction of neuronal cells in vitro. Up-regulation of SOX6 reversed the expression of miR-129-5p to reduce the damage and inflammatory response of the cell model of AD. Our study presents that up-regulation of miR-129-5p or down-regulation of SOX6 can reduce nerve injury and inflammatory response in rats with AD. Thus, miR-129-5p may be a potential candidate for the treatment of AD.

Biochemical, transgenic, and genetic complementation data demonstrate that three glutathione S-transferases are involved in the storage of anthocyanins, flavonols, and proanthocyanins in plant cells. Flavonoids are compounds in tea (Camellia sinensis) that confer the characteristic astringent taste of tea beverages; these compounds have numerous benefits for human health. In plant cells, flavonoids are synthesized in different locations within the cytoplasm and are then transported and finally stored in vacuoles. To date, the mechanism involved in the intracellular transport of flavonoids in tea has not been well elucidated. In this study, we report the functional characterization of three cDNAs encoding glutathione S-transferases (CsGSTs) of C. sinensis, namely, CsGSTa, CsGSTb, and CsGSTc. The expression profiles of CsGSTa and CsGSTb were positively correlated with the accumulation of flavonols, anthocyanins and proanthocyanins in tea tissues and cultivars. These three recombinant CsGSTs showed a high affinity for flavonols (kaempferol-3-O-glucoside and quercetin-3-O-glucoside) and anthocyanin (cyanidin-3-O-glucoside) in vitro but had no or weak affinity for epicatechin. In vivo, CsGSTa, CsGSTb and CsGSTc fully or partially restored the storage of anthocyanins and proanthocyanidins in transgenic tt19 mutants. Metabolic profiling revealed that the contents of anthocyanins, flavonols, and proanthocyanidins were increased in the transgenic petals of Nicotiana tabacum. Taken together, all data showed that CsGSTa, CsGSTb, and CsGSTc are associated with the storage of anthocyanins, flavonols, and proanthocyanins in C. sinensis cells.

Inflammatory bowel disease is a chronic nonspecific inflammatory disease of the intestine. Its pathogenesis is not yet fully understood. It may be related to heredity, environmental triggers, infection, immune dysfunction and other factors. Purinergic receptor (P2X7R) ligand-gated ion channel is closely related to inflammation and widely expressed in intestinal cells. Previous studies have shown that ATP/P2X7R signal is involved in the pathogenesis of intestinal inflammation, but its specific mechanism needs further study. This article reviews the research progress of P2X7 receptor in inflammatory bowel disease.

Atlantoaxial transarticular facet screw fixation (Magerl technique) and C1 lateral mass screws combined with C2 pedicle screws fixation (Harms technique) are the most commonly used techniques for posterior internal fixation in the upper cervical spine. Upper cervical spinal surgery is a technically demanding and challenging procedure because of complicated anatomical structures and frequent occurrence of anomalies. Accurate insertion of screws allows for stable and secure internal fixation, which is necessary for both techniques. Traditional methods under fluoroscopic assistance in this region cannot meet the requirements of high levels of accuracy and security during the procedure. Robot‐assisted spinal surgery can provide accurate and reliable guidance during the screw insertion, which is evidenced in the literature. As a recently developed technique, robot‐assisted surgery is supposed to be performed by skilled surgeons who have received standard training for robotic surgery. The standardized upper cervical spinal surgery assisted by the robot system needs to be introduced to these surgeons. Based on the consensus of consultant specialists, the literature review, and our local experience, this guideline included the introduction of the robotic system, the workflow of robot‐assisted procedures, and the precautions to take during procedures. This guideline aims to provide a standardization of the robotic surgery for posterior atlantoaxial internal fixation.

Objectives Percutaneous transfacet screw fixation (pTSF) is a minimally invasive posterior fixation technique supplementing oblique lateral interbody fusion (OLIF) for lumbar spinal disorders. Accurate screw insertion is difficult to achieve and technically demanding under 2-D fluoroscopy. Recently developed robot-assisted spinal surgery demonstrated a high level of accuracy of pedicle screw insertion and a low complication rate. No published study has reported this combination technique. The aim of our study was to evaluate the accuracy and safety properties of the combination of both minimally invasive techniques: robot-assisted pTSF supplementing the OLIF procedure. Methods This was an experimental and prospective study. Selected consecutive patients with lumbar degenerative disorders received robot-assisted pTSF supplementing the OLIF procedure using the TianJi Robot system operated by one senior surgeon from March to October 2018. The accuracy of screw insertion and perioperative screw-related complications were evaluated. Assessment of the accuracy of screw insertion included intraoperative robotic guidance accuracy and incidence of screw encroachments. Intraoperative robotic guidance accuracy referred to translational and angular deviations of screws, which were assessed by comparing the planned and actual screw trajectories guided by the robot on reconstructed images using TianJi Robot Planning Software. Screw encroachments were evaluated on postoperative CT images and classified by a grading system (A, excellent; B, good; C, poor). Screw-related complications including intraoperative pin skidding, screw malposition and adjustment, together with postoperative neurological symptoms that correlated with screw malposition were recorded. Results Ten patients, with an average age of 60.2 years, were selected and recruited in this study. All cases were degenerative lumbar spinal disorders, out of which there were 6 cases of Meyerding Grade I degenerative spondylolisthesis. Twenty-four transfacet screws were inserted by robotic assistance. Instrumented levels included nine segments at L4-5 level and three segments at L3-4 level. Two patients had both L4-5 and L3-4 level fixation. The average surgical time was 3.3 h (SD, 0.8 h). The mean blood loss was 90 mL (SD, 32 mL). Intraoperative guidance accuracy showed 1.09 ± 0.17 mm (ranging from 0.75 to 1.22 mm) translational deviation and 2.17° ± 0.39° (ranging from 1.47° to 2.54°) angular deviation. The gradings of screw encroachment were: 17 screws (71%) with Grade A, 6 screws (25%) with Grade B, and 1 screw (4%) with Grade C. Only one pin skidding occurred intraoperatively and revised subsequently. No postoperative neurological complications were found. Conclusion Our preliminary study of robot-assisted pTSF supplementing the OLIF procedure showed a high level of accuracy for screw insertion and this minimally invasive combination technique was found to be a feasible and safe procedure.

BACKGROUND: Application of tranexamic acid (TXA) in the treatment of acetabular fractures could reduce intraoperative and postoperative blood loss. OBJECTIVE: To investigate the effect of single and repeated intravenous infusion of TXA on blood loss of acetabular fractures. METHODS: 120 patients with acetabular fractures admitted to our hospital from January 2017 to September 2020 were retrospectively divided into three groups: Patients accepted 1g TXA at preoperative 30 minutes were defined as single TXA group (n = 40); Patients accepted 1g TXA at preoperative 30 minutes and 1g TXA at 3 hours after the start of surgery were defined as repeated TXA group (n = 40); Patients accepted normal saline at preoperative 30 minutes were defined as control group (n = 40). RESULTS: The total blood loss in single TXA group and repeated TXA group were significantly lower than control group, and the total blood loss in the repeated TXA group was significantly lower than single TXA group (P 0.05). CONCLUSION: Repeated TXA is more recommended during acetabular fracture surgery since it can reduce the total blood loss without increasing the operative time, postoperative transfusion rate and thrombosis rate compared with single TXA.

Excessive bleeding induces a high risk of death and is a leading cause of deaths that result from traffic accidents and military conflict. In this paper, we developed a novel porous chitosan–CaCO3 (CS–CaCO3) composite material and investigated its hemostatic properties and wound healing performance. The CS–CaCO3 composites material was prepared via a wet-granulation method. Granulation increases the infiltrating ability of the CS–CaCO3 composites material. The improved water absorption ability was enhanced to 460% for the CS–CaCO3 composites material compared to the CaCO3 or chitosan with only one single component. The coagulation studies in vivo illustrated that the blood clotting time was greatly reduced from 31 s for CaCO3 to 16 s for the CS–CaCO3 composite material. According to the results of the wound healing experiments in rats, it was found that the CS–CaCO3 composite material can promote wound healing. The CS–CaCO3 composite material could accelerate wound healing to a rate of 9 days, compared with 12 days for the CaCO3. The hemostatic activity, biocompatibility, and low cost of CS–CaCO3 composite material make it a potential agent for effective hemostatic and wound healing materials.

Background Blood transfusion can cause immunosuppression and lead to worse outcomes in patients with digestive tract malignancies; however, the specific mechanism behind this is not completely understood. One theory is that increased numbers of regulatory CD3+CD4+CD25+FOXP3+ T cells (Tregs) and forkhead box protein-3 mRNA (FOXP3) expression in the blood after transfusion contribute to these outcomes. The effect of blood transfusion on immune function in patients with different ABO blood types is variable. This study investigates the effect of intraoperative blood transfusion on the number of Tregs and the expression of FOXP3 in the blood of patients with different ABO blood types and digestive tract malignancies. Methods Patients with digestive tract malignancies who underwent radical resection and received intraoperative blood transfusion were divided into four groups according to their blood types:blood group A, blood group B, blood group O and blood group AB (n = 20 for each group). Blood was collected from all patients before surgery, immediately after transfusion, 1 day after transfusion, and 5 days after transfusion. The number of Tregs was measured by flow cytometry. The expression of FOXP3 was detected by real time reverse transcription polymerase chain reaction (RT-PCR). Results There was no significant difference in the number of Tregs or expression of FOXP3 mRNA among patients with different blood types before surgery. However, the number of Tregs and the expression of FOXP3 increased after blood transfusion in all blood type groups. This increase was especially evident and statistically significant on the first day after blood transfusion when compared with measures obtained before the surgery. Measures returned to the preoperative level five days after surgery. There were significant differences in the increase of Tregs and expression of FOXP3 among patients with different blood types. The greatest increase was seen in patients with blood group B and the least in blood group A. Conclusions Intraoperative blood transfusion can lead to an increase in blood Tregs and FOXP3 expression in patients with digestive tract malignancies. Increases were greatest on the first day after surgery and differed among patients with different blood types. Increases were greatest in blood type B and least in blood type A.

Ischemia/reperfusion (I/R)-induced rapid inflammation involving activation of leukocyte-endothelial adhesive interactions and leukocyte infiltration into tissues is a major contributor to postischemic tissue injury. However, the molecular mediators involved in this pathological process are not fully known. We have previously reported that caveolin-2 (Cav-2), a protein component of plasma membrane caveolae, regulated leukocyte infiltration in mouse lung carcinoma tumors. The goal of the current study was to examine if Cav-2 plays a role in I/R injury and associated acute leukocyte-mediated inflammation. Using a mouse small intestinal I/R model, we demonstrated that I/R downregulates Cav-2 protein levels in the small bowel. Further study using Cav-2-deficient mice revealed aggravated postischemic tissue injury determined by scoring of villi length in H&E-stained tissue sections, which correlated with increased numbers of MPO-positive tissue-infiltrating leukocytes determined by IHC staining. Intravital microscopic analysis of upstream events relative to leukocyte transmigration and tissue infiltration revealed that leukocyte-endothelial cell adhesive interactions in postcapillary venules, namely leukocyte rolling and adhesion were also enhanced in Cav-2-deficient mice. Mechanistically, Cav-2 deficiency increased plasminogen activator inhibitor-1 (PAI-1) protein levels in the intestinal tissue and a pharmacological inhibition of PAI-1 had overall greater inhibitory effect on both aggravated I/R tissue injury and enhanced leukocyte-endothelial interactions in postcapillary venules in Cav-2-deficient mice. In conclusion, our data suggest that Cav-2 protein alleviates tissue injury in response to I/R by dampening PAI-1 protein levels and thereby reducing leukocyte-endothelial adhesive interactions. NEW & NOTEWORTHY The role of caveolin-2 in regulating ischemia/reperfusion (I/R) tissue injury and the mechanisms underlying its effects are unknown. This study uses caveolin-2-deficient mouse and small intestinal I/R injury models to examine the role of caveolin-2 in the leukocyte-dependent reperfusion injury. We demonstrate for the first time that caveolin-2 plays a protective role from the I/R-induced leukocyte-dependent reperfusion injury by reducing PAI-1 protein levels in intestinal tissue and leukocyte-endothelial adhesive interactions in postcapillary venules.

STUDY DESIGN An experimental investigation of a robot-assisted ultrasonic osteotome applied to vertebral cancellous bone. OBJECTIVE The aim of this study was to investigate the effect of various ultrasonic parameter settings on temperature in the drilling site and penetration time and determine the most suitable parameters for efficient and safe robot-based ultrasonically assisted bone drilling in spinal surgery. SUMMARY OF BACKGROUND DATA A robot-assisted ultrasonic osteotome device may be safe and effective for spinal drilling. METHODS Sixty specimens of bovine vertebral cancellous were randomly assigned to one of six groups, which varied by mode of ultrasonic vibration (L-T and L) and feed rate (one percent [0.8 mm/s], two percent [1.6 mm/s], and three pecent [2.4 mm/s]). Maximum temperature in the drilling site and penetration time was recorded. RESULTS Maximum temperature in the drilling site decreased as output power increased for L-T and L modes, was significantly lower for L-T compared to L mode at each feed rate and power setting, was significantly different at feed rates of 1.6 mm/s versus 0.8 mm/s and 2.4 mm/s versus 0.8 mm/s for L-T mode at an output power of 60 W and 84 W, but was not influenced by feed rate for L mode. Penetration time did not significantly improve as output power increased for both L-T and L modes, was significantly decreased with increased feed rates, but was not significantly different between L-T and L modes. CONCLUSION The optimal parameters for applying a robot-assisted ultrasonic osteotome to vertebral cancellous bone are L-T mode, maximum output power of 120 W, and maximum feed rate of 2.4 mm/s.Level of Evidence: 4.

piRNAs are a class of small non-coding RNA molecules, which interact with the PIWI family and have many important and diverse biological functions. The present review is aimed to provide guidelines and contribute to piRNA research. We focused on the four types of identification models on piRNA-related molecules, including piRNA, piRNA cluster, piRNA target, and disease-related piRNA. We evaluated the types of tools for the identification of piRNAs based on five aspects: datasets, features, classifiers, performance, and usability. We found the precision of 2lpiRNApred was the highest in datasets of model organisms, piRNN had a better performance of datasets of non-model organisms, and 2L-piRNA had the fastest recognition speed of all tools. In addition, we presented an overview of piRNA databases. The databases were divided into six categories: basic annotation, comprehensive annotation, isoform, cluster, target, and disease. We found that piRNA data of non-model organisms, piRNA target data, and piRNA-disease-associated data should be strengthened. Our review might assist researchers in selecting appropriate tools or datasets for their studies, reveal potential problems and shed light on future bioinformatics studies.

Although malignant glioblastoma (GBM) treatment has significantly improved in the past few decades, the prognosis of GBM remains unsatisfactory. MicroRNA (miR)-138-5p has been reported as a tumor suppressor in several types of human cancer; however, little is known about the function of miR-138-5p in GBM. The present study aimed to investigate the role of miR-138-5p in GBM as well as the underlying molecular mechanisms. The present study performed bioinformatics analysis, reverse transcription-quantitative (RT-q)PCR, western blotting, cell viability assays, colony formation assays, invasion assays and cell cycle analysis to investigate the biological function of miR-138-5p in both patient tissues and cell lines. In addition, miR-138-5p targets in GBM were predicted using Gene Expression Omnibus website and further validated by a dual luciferase reporter gene assay. The results revealed that miR-138-5p expression levels in patients with GBM from a Gene Expression Omnibus dataset were significantly downregulated. RT-qPCR analysis of miR-138-5p expression levels also revealed similar results in GBM tissues and cell lines. The upregulation of miR-138-5p expression levels using a mimic significantly inhibited the cell viability, colony formation and the G0/G1 to S progression in GBM cell lines, suggesting that miR-138-5p may be a tumor suppressor. Moreover, miR-138-5p was discovered to directly target cyclin D3 (CCND3), a protein that serves an important role in the cell cycle, and inhibited its expression. Finally, silencing CCND3 using small interfering RNA suppressed the viability of GBM cells. In conclusion, the results of the present study suggested that miR-138-5p may function as a tumor suppressor in GBM by targeting CCND3, indicating that miR-138-5p may be a novel therapeutic target for patients with GBM.

Objective Current treatment and prognosis of Parkinson’s disease (PD) are not ideal. This study explored the mechanism of long non-coding RNA (lncRNA) rhabdomyosarcoma 2-associated transcript (RMST) in dopaminergic (DA) neuron damage in PD rats. Methods PD rats were modeled and injected with RMST silence or overexpression vectors to figure out its roles in oxidative stress, the apoptosis of DA neurons in brain substantia nigra (SN), and neurobehavioral activities of PD rats. Tyrosine hydroxylase (TH), synaptophysin (SYN), glial fibrillary acidic protein (GFAP) and ionized calcium-binding adaptor molecule (Iba-1) in SN were detected. RMST and Toll-like receptor (TLR)/nuclear factor kappa B (NF-κB) pathway-related factors were detected. Results RMST expression in brain SN of rats, TLR2, TLR4 expression in neurons and NF-κB expression in cell nucleus were increased. Silenced RMST improved the neurobehavioral activities, depressed oxidative stress and neuronal apoptosis, increased TH and SYN expression, and reduced the activation degree of glial cells in SN and the inflammatory response via reducing GFAP and Iba-1. Moreover, reduced RMST reduced TLR2 and TLR4 expression in neurons and NF-κB expression in cell nucleus in PD rats. Conclusion Inhibited RMST attenuates DA neuron damage in PD rats, which may be implicated with TLR/NF-κB signaling pathway.

Background Acute myeloid leukemia is a genetically, biologically and clinically heterogeneous hematopoietic malignancy that is highly dependent on the bone marrow microenvironment. Infiltrated immune cells and stromal cells are an important part of the bone marrow microenvironment and significantly affect the progression of acute myeloid leukemia. Recently, the competing endogenous RNA hypothesis has gained great interests in the study of molecular and biological mechanisms of tumor occurrence and progression. However, research on how competing endogenous RNA relates to leukemia tumor microenvironment remains uninvestigated.Methods In this study, mRNA, miRNA and lncRNA data and clinical information of the acute myeloid leukemia cohort were obtained from The Cancer Genome Atlas database, and the immune and stromal scores were calculated using the ESTIMATE algorithm.Results We found that immune scores were significantly correlated with cytogenetic risk and overall survival, and also identified microenvironment-related mRNAs, miRNAs, and lncRNAs based on the immune and stromal scores. Differentially expressed mRNAs and lncRNAs were applied to weighted correlation network analysis to identify the modules most relevant to the immune microenvironment of acute myeloid leukemia. Using miRNA database to predict miRNA-targeted genes, we established the immune-related competing endogenous RNA network consisting of 33 lncRNAs, 21 miRNAs and 135 mRNAs. Prognostic analysis was performed on the genes in the immune-related competing endogenous RNA network to screen out 15 lncRNAs, 2 miRNAs and 31 mRNAs with prognostic values.Conclusions In summary, we identified a novel immune-related mRNA-miRNA-lncRNA competing endogenous RNA network associated with the prognosis of acute myeloid leukemia, which may contribute to better understanding of the development and progression of acute myeloid leukemia and to serve as novel therapeutic targets.

Inflammatory bowel diseases (IBDs) are chronic immunological disorders of the intestinal tract characterized by persistent inflammation. Baicalin, a type of flavonoid, has exhibited a wide range of pharmacological activities, including immunomodulation and anti-inflammation. However, little is known about the therapeutic role of baicalin in IBD. The aim of the present study was to ascertain whether baicalin could be a therapeutic drug of IBD and investigate its specific mechanisms. In the present study, the results revealed that baicalin not only significantly alleviated TNBS-induced colitis by reducing the release of IL-6, TNF-α and IL-1β and increasing the level of IL-10, but promoted the expression of tight-junction proteins ZO-1 and β-catenin, which may have been achieved by blockage of the PI3K/AKT signaling pathway. In vitro, the results demonstrated that baicalin clearly inhibited the release of TNF-α, IL-6 and IL-1β and promoted the expression of IL-10 in LPS-induced HT-29 cells, and significantly decreased LPS-induced HT-29 cell apoptosis by blockage of the PI3K/AKT signaling pathway. In conclusion, the present research revealed for the first time that baicalin acted as a therapeutic drug in IBD by suppression of the PI3K/AKT signaling pathway.

The role of genomic variation in differentiation is currently not well understood. Here, the genomic variations were determined with the whole-genome sequencing for three pairs of pluripotent stem cell lines and their corresponding BMP4-induced trophoblast cell lines. We identified ∼3,500 single nucleotide variations and ∼4,500 indels by comparing the genome sequenced data between the stem cell lines and the matched BMP4-induced trophoblast cell lines and annotated them by integrating the epigenomic and transcriptomic datasets. Relatively, introns enrich more variations. We found ∼45% (42 genes) of the differentially expressed genes in trophoblasts that associate genomic variations. Six variations, located at transcription factor binding sites where H3K4me3 and H3K27ac are enriched in both H1 and H1_BMP4, were identified. The epigenetic status around the genomic variations in H1 was similar to that in H1_BMP4. This means that the variation-associated gene’s expression change can not be attributed to epigenetic alteration. The genes associated with the six variations were upregulated in differentiation. We inferred that during the differentiation, an increased in the expression level of the MEF2C gene is due to a genomic variation in chromosomes 5: 88179358 A > G, which is at a binding site of TFs KLF16, NR2C2, and ZNF740 to MEF2C. Allele G shows a higher affinity to the TFs in the induced cells. The increased expression of MEF2C leads to an increased expression of TF MEF2C’s target genes, subsequently affecting the differentiation. Although genomic variation should not be a dominant factor in differentiation, we believe that genomic variation could indeed play a role in the differentiation from stem cells into trophoblast.

Objective Parkinson’s disease (PD) is a frequent degenerative disease of the nervous system with undefined pathogenesis. This study explored the protective effect of microRNA (miR)-218−5p on dopaminergic neuron injury in substantia nigra (SN) of rats with PD through the regulation of LIM and SH3 domain protein 1 (LASP1). Methods The PD rat model was established by fixed point injection of 6-hydroxydopamine into the rats. The PD rats were injected with miR-218−5p overexpressed recombinant adeno-associated virus (rAAV) or LASP1 silenced rAAV to explore their roles in dopaminergic neurons in SN of rats with PD. The changes in pathological structure of SN were observed and the expression of tyrosine hydroxylase (TH) and deacetylvindoline acetyltransferase (DAT), the dopaminergic neuron apoptosis and oxidative stress factor in the SN were detected. The expression of miR-218−5p, LASP1, Bcl-2 and Bax in SN was detected. The targeting relationship between miR-218−5p and LASP1 was confirmed. Results Declined miR-218−5p and overexpressed LASP1 existed in the brain SN of PD rats. Up-regulated miR-218−5p or inhibited LASP1 improved the pathological damage of dopaminergic neurons and increased the number of TH and DAT positive cells in brain SN of PD rats. Furthermore, elevated miR-218−5p or depressed LASP1 inhibited the apoptosis, and oxidative stress of dopaminergic neurons in brain SN of PD rats. In addition, increased miR-218−5p repressed the expression of LASP1 in the brain SN of PD rats. LASP1 was proven to be a direct target of miR-218−5p. Conclusion The study highlights that up-regulated miR-218−5p could improve the damage of dopaminergic neurons in PD rats, which was related to the inhibition of LASP1.

AIMS To investigate the association of several single nucleotide polymorphisms (SNPs) within alpha-synuclein (SNCA) gene and additional gene-environment interaction with Parkinson's disease (PD) risk. METHODS Hardy-Weinberg equilibrium (HWE) is tested for controls using SNPstats (http://bioinfo.iconcologia.net/SNPstats). Logistic regression is used to calculate the ORs (95% CI) for relations between the four SNPs and PD risk. The generalized multifactor dimensionality reduction (GMDR) model is used to evaluate the synergy between gene and environment. RESULTS A total of 1161 people were included in this study, including 386 cases of PD and 775 normal controls. In this study, the genotype frequency of the control group was consistent with HWE distribution. Rs356219-G allele frequency was 30.0% in patients and 19.8% in control group. The rs356221-T allele frequency was 29.7% in the patients and 20.8% in the control group. Rs356219-G and rs356221-T alleles were associated with increased PD risk, with adjusted ORs (95% CI) of 1.92 (1.28-2.52) and 1.52 (1.05-2.02), respectively. We also found no significant correlation between rs2301134 and rs2301135 and susceptibility to PD. The best gene-environment interaction models were determined by GMDR analysis, which shown a significant gene-T2DM interaction combinations, but the gene-alcohol drinking interaction combinations were all not significant. We also conducted stratified analysis for interaction effect using logistic regression. We found that T2DM patients with rs356221-AT/ TT genotype have the highest PD risk, compared to subjects with rs356219-AA genotype, OR (95%CI) = 2.67 (1.83-3.46). CONCLUSIONS The rs356219-G and rs356221-T, gene-environment interaction between rs356221 and T2DM were all associated with increased PD risk.

Study design A model experiment. Objective To measure and compare the clinical accuracy of point-to-point registration (PR) and auto-registration (AR) in an operative set using an active infrared navigation system. Summary of background data PR and AR are two major registration methods of navigation assisted spinal surgery. No previous study compared the difference between the two methods with respect to clinical accuracy. Methods A novel method was used to measure the clinical accuracy of the navigation system under an operative set using a Sawbone model with titanium beads on the surface, which was essential to measure the accuracy numerically, instead of a real patient. Both the operative set and the procedure mimicked a regular surgery. The clinical accuracy was defined as the average distance between the "navigation coordinate" and the "image coordinate." The clinical accuracy of the PR using preoperative computed tomography (CT) images and the AR using intraoperative CT images was measured and compared. Results The average clinical accuracy of PR was different among different segments. The accuracy of the most accurate segment, which provided the reference points during the PR, was 1.10 mm. In the two segments adjacent to the reference segment, the clinical accuracy deteriorated to 1.37 and 1.50 mm. The accuracy of the farther segments was worse. In comparison, the clinical accuracy of different segments of AR was of no significant difference. The average accuracy of AR was 0.74 mm, which was significantly better than the best accuracy of PR. Conclusion AR is better than PR with respect to clinical accuracy in navigation assisted spinal surgery. Level of evidence N/A.

Tea possesses a distinctive flavor profile and can have health benefits owing to the high levels of flavonoids in its leaves. However, the mechanism of the flavonoid glycosylation hasn't been well studied in tea plants, especially glycosylation at the 7-OH site has rarely been reported. In this study, four UGT genes CsUGT73A20, CsUGT75L12, CsUGT78A14 and CsUGT78A15 were isolated from tea leaves and overexpressed in the model plants Arabidopsis thaliana and Nicotiana tabacum for the functional identification of genes in vivo. In order to characterize the CsUGT functions in model plants, flavonoids in seeds of Arabidopsis and the flowers of tobacco were identified first. In CsUGT73A20-overexpressing Arabidopsis and tobacco, the level of certain flavonol glycosides involved in glycosylation reactions at the 3-OH and 7-OH sites increased considerably, but the level of flavan-3-ols decreased. In CsUGT75L12 transgenic Arabidopsis, the level of flavonol glycosides exhibiting glucosyltransferase activity at the 7-OH position increased markedly, but the concentrations of quercetin and kaempferol and flavan-3-ols decreased. In both transgenic Arabidopsis and tobacco, CsUGT78A14 promoted the synthesis of more flavonol glucosides with UDP-glucose as a sugar donor at the 3-OH glycosylation site. In CsUGT78A15 transgenic plants, flavonol galactosides at the 3-OH glycosylation site with UDP-galactose as a sugar donor were increased. In the tea plant, the corresponding flavonoid glycosides such as kaempferol‑3‑O‑β‑d‑glucosides, kaempferol‑3‑O‑β‑d‑galactosides, kaempferol‑7‑O‑β‑d‑glucoside, and luteolin‑7‑O‑β‑d‑glucoside were identified. And it could be possible that they were products of CsUGT78A14, CsUGT78A15, CsUGT73A20 and CsUGT75L12, respectively.

Background Cervical spondylosis adversely affects life quality for its heavy disease burden. The report on the community-based prevalence and associated factors of cervical spondylosis is rare, especially in Chinese population. Whether prevention is needed and how to prevent it is not clear. This study aims to explore its prevalence and related lifestyle factors and provide evidence on prevention of cervical spondylosis. Methods A community-based multistage cross-sectional survey of six communities from the Chinese population was conducted. A face-to-face interview was conducted to obtain individual information, and prevalence was calculated. Single-factor analysis and multivariable logistic regressions were used to explore the associated factors in total and subgroup populations. Results A total of 3859 adults were analyzed. The prevalence of cervical spondylosis was 13.76%, although it differed significantly among the urban, suburban, and rural populations (13.07%, 15.97%, and 12.25%, respectively). Moreover, it was higher in females than in males (16.51% vs 10.49%). The prevalence among different age groups had an inverted U shape. The highest prevalence was in the age group from 45 to 60 years old. The associated factors differed by subgroups. There were positive associations between engaging in mental work, high housework intensity, and sleep duration of less than 7 h/day with cervical spondylosis. Going to work on foot was a negative factor of cervical spondylosis in the total population. For people aged less than 30 years, keeping the same work posture for 1–2.9 h/day was a special related factor. Exposure to vibration was an associated factor for females aged 45–60 years. Menopause was a special related factor for women. Conclusions Prevalence of cervical spondylosis was high in Chinese population. People younger than 60 years were the focus of prevention for cervical spondylosis. Moreover, the characters between male and female and among different age groups were different and required targeted interventions.

Carbon black (CB), a core elemental carbon component of airborne particles, has been used as a model material to study environmental safety and health impacts of airborne particles. Although potential adverse effects of CB have been reported, limited knowledge is available regarding CB-induced metabolic disorders and secondary effects distant from primary target organs, such as the effects on joints. The knee cavity is a relatively closed space along the peripheral circulation route with a slow rate of interchange of nutrition with blood. While epidemiologic studies have indicated that airborne particle exposure may affect the occurrence and severity of inflammatory knee diseases, no research has been performed to understand the potential hazardous direct/indirect interactions between particles and knee cells. Herein, we have scrutinized the toxicity of four commercial nano-sized CB samples in the lung and a distant site: knee joint. Our results indicated that CB triggered pulmonary and systemic inflammation upon inhalation exposure, and, more strikingly, CB also elicited injuries of the knee joint, as demonstrated by thickened synovial membrane, suggesting disordered cellular metabolism within the knee joint. Our data recognized the CB toxicity profiles to macrophages as characterized by pro-inflammatory reactions, and also defined an activated metabolic state of chondrocytes, as evidenced by metalloproteinase (MMP) induction. Of note, remarkable variations were also found for these changes induced by these four CB samples, due to their distinct physicochemical properties. Collectively, our results uncovered a significant toxicity of CB inhalation exposure to the knee joint, as reflected by metabolic activation of chondrocytes, and, more importantly, these findings unearthed CB-induced metabolic disorders and secondary effects owing to systemic pro-inflammatory conditions upon CB exposure, in addition to the likelihood of direct toxicity to knee cells.

Tea is an important economic crop with a 3.02 Gb genome. It accumulates various bioactive compounds, especially catechins, which are closely associated with tea flavor and quality. Catechins are biosynthesized through the phenylpropanoid and flavonoid pathways, with 12 structural genes being involved in their synthesis. However, we found that in Camellia sinensis the understanding of the basic profile of catechins biosynthesis is still unclear. The gene structure, locus, transcript number, transcriptional variation, and function of multigene families have not yet been clarified. Our previous studies demonstrated that the accumulation of flavonoids in tea is species, tissue, and induction specific, which indicates that gene coexpression patterns may be involved in tea catechins and flavonoids biosynthesis. In this paper, we screened candidate genes of multigene families involved in the phenylpropanoid and flavonoid pathways based on an analysis of genome and transcriptome sequence data. The authenticity of candidate genes was verified by PCR cloning, and their function was validated by reverse genetic methods. In the present study, 36 genes from 12 gene families were identified and were accessed in the NCBI database. During this process, some intron retention events of the CsCHI and CsDFR genes were found. Furthermore, the transcriptome sequencing of various tea tissues and subcellular location assays revealed coexpression and colocalization patterns. The correlation analysis showed that CsCHIc, CsF3'H, and CsANRb expression levels are associated significantly with the concentration of soluble PA as well as the expression levels of CsPALc and CsPALf with the concentration of insoluble PA. This work provides insights into catechins metabolism in tea and provides a foundation for future studies.

piwi-interacting RNAs (piRNAs) are valuable biomarkers, but functional studies are still very limited. Recent research shows that piRNA-mediated cleavage acts on Transposable Elements (TEs), messenger RNAs (mRNAs), and long non-coding RNAs (lncRNAs). This study aimed to predict cancer-associated piRNA-mRNA and piRNA-lncRNA interactions as well as piRNA regulatory functions. Four cancer types (BRCA, HNSC, KIRC, and LUAD) were investigated. Interactions were identified by integrated analysis of the expression and sequence data. For the expression analysis, only piRNA-mRNA and piRNA-lncRNA pairs with expression profiles that were significantly inversely correlated were retained to reduce false-positive rates during the prediction. For the sequence analysis, miRanda was used for the target prediction. We identified 198 piRNA-mRNA and 10 piRNA-lncRNA pairs. Unlike mRNA and lncRNA expressions, the piRNA expression was relatively consistent across the cancer types. Furthermore, the identified piRNAs were consistent with previously published cancer biomarkers, such as piRNA-36741, piR-21032, and piRNA-57125. More importantly, predicted piRNA functions were determined by constructing an interaction network, and piRNA targets were placed in gene ontology categories related to the cancer hallmarks “activating invasion and metastasis” and “sustained angiogenesis”.

Ursolic acid (UA), a natural triterpene acid, is a promising anti-cancer drug due to its inhibitory effect on tumor growth and metastasis. However, clinical translation of UA is limited by its poor water solubility and low bioavailability. To overcome these problems, herein an amphiphilic self-assembly nanodrug composed of UA, lactobionic acid (LA) and low-polyamidoamine (low-PAMAM) dendrimers is developed. This near-spherical nanodrug with a uniform size (~180 nm) demonstrated to have an enhanced cytotoxicity against liver cancer SMMC7721 cells, and could attenuate the migration and adhesion of SMMC7721 cells at non-toxic concentrations by suppressing metastasis-related protein MMP-9 expression. Furthermore, in vivo study indicates that the nanodrug exhibited prolonged circulation time in blood as well as increased AUC, MRT and Cmax, and could effectively inhibit the tumor growth in H22 mice model. Overall, the UA-based nanodrug delivery system reported in the present work represents a novel strategy for targeted tumor therapy.

There were several high concentrations of flavonoid components in tea leaves that present health benefits. A novel purple-leaf tea variety, ‘Mooma1’, was obtained from the natural hybrid population of Longjing 43 variety. The buds and young leaves of ‘Mooma1’ were displayed in bright red. HPLC and LC-MS analysis showed that anthocyanins and O-Glycosylated flavonols were remarkably accumulated in the leaves of ‘Mooma1’, while the total amount of catechins in purple-leaf leaves was slightly decreased compared with the control. A R2R3-MYB transcription factor (CsMYB6A) and a novel UGT gene (CsUGT72AM1), that were highly expressed in purple leaf were isolated and identified by transcriptome sequencing. The over-expression of transgenic tobacco confirmed that CsMYB6A can activate the expression of flavonoid-related structural genes, especially CHS and 3GT, controlling the accumulation of anthocyanins in the leaf of transgenic tobacco. Enzymatic assays in vitro confirmed that CsUGT72AM1 has catalytic activity as a flavonol 3-O-glucosyltransferase, and displayed broad substrate specificity. The results were useful for further elucidating the molecular mechanisms of the flavonoid metabolic fluxes in the tea plant.

Polyphenols are one of the largest groups of compounds that confer benefits to the health of plants and humans. Flavonol glycosides are a major ingredient of polyphenols in Camellia sinensis. Flavonol-3-O-glycosides are characteristic astringent taste compounds in tea infusion. A polyphenolic glycosyltransferase (CsUGT72AM1) belonging to cluster IIIb was isolated from the tea plant. The full-length cDNA of CsUGT72AM1 is 1416 bp. It encodes 472 amino acids with a calculated molecular mass of 50.92 kDa and an isoelectric point of 5.21. The recombinant CsUGT72AM1 protein was expressed in Escherichia coli and exhibited catalytic activity toward multiple flavonoids and coniferyl aldehyde. The enzyme assay indicated that rCsUGT72AM1 could perform glycosidation of flavonols or coniferyl aldehyde in vitro to form 3-O-glucoside or 4-O-glucoside, respectively. Interestingly, this enzyme also had activities and performed multisite glycosidation toward flavanones. The consistent products were confirmed to be naringenin-7-O-glucoside and -4'-O-glucoside by the nuclear magnetism assay. In addition, in the enzyme assay with cyanidin as the substrate, the results suggested that the glycosylated activity of CsUGT72AM1 was remarkably inhibited by a high concentration of anthocyanins. The above results indicate that CsUGT72AM1 may be involved in the metabolism of flavonol, flavanone, anthocyanin, and lignin.

Activation of large-conductance Ca2+-activated K+ (BKCa) channels evokes cell survival programs that mitigate intestinal ischemia and reperfusion (I/R) inflammation and injury 24 h later. The goal of the present study was to determine the roles of reactive oxygen species (ROS) and heme oxygenase (HO)-1 in delayed acquisition of tolerance to I/R induced by pretreatment with the BKCa channel opener NS-1619. Superior mesentery arteries were occluded for 45 min followed by reperfusion for 70 min in wild-type (WT) or HO-1-null (HO-1−/−) mice that were pretreated with NS-1619 or saline vehicle 24 h earlier. Intravital microscopy was used to quantify the numbers of rolling and adherent leukocytes. Mucosal permeability, tumor necrosis factor-α (TNF-α) levels, and HO-1 activity and expression in jejunum were also determined. I/R induced leukocyte rolling and adhesion, increased intestinal TNF-α levels, and enhanced mucosal permeability in WT mice, effects that were largely abolished by pretreatment with NS-1619. The anti-inflammatory and mucosal permeability-sparing effects of NS-1619 were prevented by coincident treatment with the HO-1 inhibitor tin protoporphyrin-IX or a cell-permeant SOD mimetic, Mn(III)tetrakis (4-benzoic acid) porphyrin (MnTBAP), in WT mice. NS-1619 also increased jejunal HO-1 activity in WT animals, an effect that was attenuated by treatment with the BKCa channel antagonist paxilline or MnTBAP. I/R also increased postischemic leukocyte rolling and adhesion and intestinal TNF-α levels in HO-1−/− mice to levels comparable to those noted in WT animals. However, NS-1619 was ineffective in preventing these effects in HO-1-deficient mice. In summary, our data indicate that NS-1619 induces the development of an anti-inflammatory phenotype and mitigates postischemic mucosal barrier disruption in the small intestine by a mechanism that may involve ROS-dependent HO-1 activity. NEW & NOTEWORTHY Antecedent treatment with the large-conductance Ca2+-activated K+ channel opener NS-1619 24 h before ischemia-reperfusion limits postischemic tissue injury by an oxidant-dependent mechanism. The present study shows that NS-1619-induced oxidant production prevents ischemia-reperfusion-induced inflammation and mucosal barrier disruption in the small intestine by provoking increases in heme oxygenase-1 activity.

Genome-wide association study is especially challenging in detecting high-order disease-causing models due to model diversity, possible low or even no marginal effect of the model, and extraordinary search and computations. In this paper, we propose a niche harmony search algorithm where joint entropy is utilized as a heuristic factor to guide the search for low or no marginal effect model, and two computationally lightweight scores are selected to evaluate and adapt to diverse of disease models. In order to obtain all possible suspected pathogenic models, niche technique merges with HS, which serves as a taboo region to avoid HS trapping into local search. From the resultant set of candidate SNP-combinations, we use G-test statistic for testing true positives. Experiments were performed on twenty typical simulation datasets in which 12 models are with marginal effect and eight ones are with no marginal effect. Our results indicate that the proposed algorithm has very high detection power for searching suspected disease models in the first stage and it is superior to some typical existing approaches in both detection power and CPU runtime for all these datasets. Application to age-related macular degeneration (AMD) demonstrates our method is promising in detecting high-order disease-causing models.

Drug nanocarriers that have long been expected to revolutionize cancer therapy have yet to achieve the significant clinical success. Therefore, it remains controversial to pursue a complex drug nanocarrier that lacks clinical relevance. Herein, we developed an easily-synthesized anti-tumor drug that actively seeks the acidic tumor microenvironment while ignoring the normal tissue without the aid of additional carriers. This called "carrier-free" drug (CFD) is capable of switching its morphology from the unstructured solution to the spherical structure in response to tumor acidity. CFDs were the water-soluble zwitterionic unimers to prevent the non-specific distribution in the circulation, whereas they spontaneously formed into the particles tending to accumulation in tumor. CFD overloading in tumor cells caused the lysosomal dysfunction and autophagy blockage, thereby triggered the cell death. All the in vitro and in vivo data demonstrated the tumor-acidity-selective cytotoxicity of CFD. This facile strategy to create a self-delivering anticancer drug may cast a new light on the development of cancer therapy.

This study was a retrospective review of prospectively collected clinical data.To evaluate the clinical and radiologic outcomes of computer-assisted minimally invasive spine surgery transforaminal lumbar interbody fusion (CAMISS-TLIF) and open TLIF for the treatment of 1-level degenerative lumbar disease.Minimally invasive TLIF is becoming increasingly popular; however, the limited space and high rate of hardware complications associated with this method are challenging to surgeons. Computer-assisted navigation has the potential to dynamically show the fine anatomic structures, which could theoretically facilitate minimally invasive spine procedures.Sixty-one patients underwent 1-level TLIF procedures (30, CAMISS-TLIF; 31, open TLIF). The computer-assisted navigation system was used for CAMISS-TLIF, whereas conventional fluoroscopy was used for open TLIF. Demographic, operative, visual analog scale, and Oswestry disability index data were collected. Screw insertion was assessed by computed tomography, and radiologic fusion based on Bridwell grading was evaluated 2 years after surgery by independent investigators.The CAMISS-TLIF group had significantly less blood loss, postoperative drain, need for transfusion, and initial postoperative back pain; earlier rehabilitation; and shorter postoperative hospitalization than the open TLIF group, whereas CAMISS-TLIF took longer surgical time than open TLIF. However, no significant differences between the 2 groups in visual analog scale scores and Oswestry disability index were observed at 3 months, 1 year, and 2 years postoperatively. A total of 93.33% and 73.39% of screws in the CAMISS and open groups, respectively, had no pedicle perforation (P=0.016), and the fusion rate was similar in both groups (P=0.787).Computer-assisted navigation facilitated minimally invasive spine surgery-TLIF. CAMISS-TLIF was superior to open TLIF for treating 1-level degenerative lumbar disease, although it required longer operation time in the initial stage. CAMISS-TLIF showed several benefits compared with open TLIF, including less intraoperative blood loss, postoperative drainage, and pain; earlier rehabilitation; and shorter postoperative hospitalization.

Phenolic compounds, such as flavanols, flavones, flavonols, anthocyanins, and phenolic acids, contribute to a plant’s adaptation to their ecological niches. These compounds serve as attractants of organisms beneficial to the plants or as defence against other biotic and abiotic factors during the plant evolution. In this paper, the accumulation patterns of phenolic compounds in the eight evergreen woody core eudicots indicated divergent and convergent evolution of plant secondary metabolites. The results indicated some phenolic acids and flavonols were distributed in all the studied eight evergreen woody plants, within Oleaceae, Theaceae, Caprifoliaceae and Celastraceae, with different accumulation profiles. The flavan-3-ols, monomeric catechins, and polymeric proanthocyanidins, were undetectable in Osmanthus fragrans, Ligustrum lucidum, and Fraxinus mariesii of Oleaceae, but highly accumulated in the other five species, especially in Camellia sinensis of Theaceae. It indicated Camellia sinensis evolved great ecological adaptive capacity to prevent herbivores, insect pests, and microbial pathogens.

Background/Aims: The study aimed to investigate the protective effect of curcumin against oxidative stress-induced injury of Parkinson’s disease (PD) through the Wnt/β-catenin signaling pathway in rats. Methods: The successfully established PD rat models and normal healthy rats were randomly assigned into the 6-hydroxydopamine (6-OHDA), the curcumin (Cur) and the control groups. Immunohistochemistry was used to detect the positive expression of tyrosine hydroxylase (TH), dopamine transporter (DAT) and glial fibrillary acidic protein (GFAP). Deutocerebrum primary cells were extracted and classified into the control, 6-OHDA, Cur (5, 10, 15 µmol/L), Dickkopf-1 (DKK-1) and Cur + DKK-1 groups. MTT assays, adhesion tests and TUNEL staining were used to assess cell viability, adhesion and apoptosis, respectively. Western blotting and qRT-PCR were used to examine the protein and mRNA expressions of Wnt3a and β-catenin and the c-myc and cyclinD1 mRNA expressions. Results: TH and DAT expressions in the Cur group were elevated and GFAP was reduced compared with the 6-OHDA group. Curcumin enhanced viability, survival and adhesion and attenuated apoptosis of deutocerebrum primary cells by activating the Wnt/β-catenin signaling pathway. Higher Wnt3a and β-catenin mRNA and protein expressions and c-myc and cyclinD1 mRNA expressions, enhanced superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) contents, decreased malondialdehyde (MDA) content and elevated mitochondrial membrane potential (∆ψm) were found in the 10 and 15 µmol/L Cur groups compared with the 6-OHDA group. However, opposite tendencies were found in the Cur + DKK-1 group compared to the 10 µmol/L Cur group. Conclusion: This study suggests that curcumin could protect against oxidative stress-induced injury in PD rats via the Wnt/β-catenin signaling pathway.

Ursolic acid (UA) has been recently used as a promising anti-tumor and cancer metastatic chemo-preventive agent due to its low toxicity and liver-protecting property. However, the low bioavailability and nonspecific tumor targeting restrict its further clinical application. To address the problem, a silica-based mesoporous nanosphere (MSN) controlled-release drug delivery system (denoted UA@M-CS-FA) was designed and successfully synthesized, and was functionalized with folic acid (FA) and pH-sensitive chitosan (CS) for the targeted delivery of UA to folate receptor (FR) positive tumor cells. UA@M-CS-FA were spherical with mean diameter below 150 nm, and showed about -20 mV potential. Meanwhile, UA@M-CS-FA exhibited a pH-sensitive release manner and high cellular uptake in FR over-expressing HeLa cancer cells. Also, in vitro cellular assays suggested that UA@M-CS-FA inhibited cancer cell growth, invasion and migration. Mechanistically, UA@M-CS-FA induced cancer cell apoptosis and inhibited migration via cell cycle arrest in the G0/G1 stage, regulating the PARP/Bcl-2/MMP-9/CD44/PTEN/P53. Importantly, in vivo experiments further confirmed that UA@M-CS-FA significantly suppressed the tumor progression and lung metastasis in tumor-bearing nude mice. Immunohistochemical analysis revealed that UA@M-CS-FA treatment regulated CD44, a biomarker of cancer metastasis. Overall, our data demonstrated that a CS and FA modified MSN controlled-release drug delivery system could help broaden the usage of UA and reflect the great application potential of the UA as an anticancer or cancer metastatic chemopreventive agent.

Ursolic acid (UA) as a nature product exhibits good anti-cancer activity, low toxicity, and good liver protection features. However, the low-solubility and poor bioavailability restrict its further clinical application. To overcome this problem, a pH-sensitive prodrug delivery system (UA@MSN-UA) that incorporated acid-sensitive linkage between drug and silica-based mesoporous nanosphere (MSN) was successfully designed and synthesized. The physicochemical properties of the UA@MSN-UA nanoparticles were investigated for shape, particle size, zeta potential, nitrogen adsorption-desorption and infrared (IR) spectroscopy. The nanoparticles were further evaluated for in vitro cytotoxicity, including proliferation inhibition, cell cycle distribution and apoptotic effects against human hepatocellular carcinoma HepG2 cells. The TEM image showed that the size of synthesized MSN nanoparticle was a near-spherical shape with ~100nm diameter. In vitro cytotoxicity testing demonstrated that UA@MSN-UA nanoparticles prodrug exhibited higher proliferation inhibition, cell cycle arrest at the G2/M phase and significantly caused the early and late apoptosis in HepG2 cells, which would be contributed to high loading capacity, high cellular uptake and sustained release of UA. Overall, the UA-modified MSN prodrug delivery system can be a promising drug carrier for improving the bioavailability of UA, and further enhance its anti-cancer efficacy.

Anthocyanins are plant-specific pigments, the biosynthesis of which is stimulated by pathogen infection in several plant species. A. thaliana seedlings injected with airborne fungi can accumulate a high content of anthocyanins. The mechanism involved in fungus-induced anthocyanin accumulation in plants has not been fully described. In this study, the fungus Penicillium corylophilum (P. corylophilum), isolated from an Arabidopsis culture chamber, triggered jasmonic acid (JA), salicylic acid (SA), and anthocyanin accumulation in A. thaliana. Inhibitors of JA and SA biosynthesis suppressed the anthocyanin accumulation induced by P. corylophilum. The anthocyanin content was minimal in both the null mutant of JA-receptor coi1 and the null mutant of SA-receptor npr1 under P. corylophilum stimulation. The results indicate that JA and SA signaling mediated fungus-induced anthocyanin biosynthesis in A. thaliana. P. corylophilum led to different levels of anthocyanin generation in null mutants for MYB75, bHLH, EGL3, and GL3 transcription factors and WD40 protein, demonstrating that multiple MYB-bHLH-WD40 transcription factor complexes participated in fungus-induced anthocyanin accumulation in A. thaliana. The present study will help further elucidate the mechanism of plant resistance to pathogen infection.

Wild tea plants, which are classified into different species in the section Thea of the genus Camellia, are widely distributed in southern China. Tea produced from these plants has a unique flavor, which is different from that of tea produced from tea cultivars. In this study, we performed a comparative analysis of morphology, phylogenetic relationships, and phenolic compound metabolism between two wild tea plants (Gujing and Siqiu) and a tea cultivar (Shuchazao). Siqiu and Gujing tea plants had similar morphological traits and could be phylogenetically classified into a same cluster, which was entirely separate from the cluster containing widely cultivated cultivars such as Camellia sinensis cv. Shuchazao. Combined metabolomic and transcriptome analyses revealed that UGT84a22 was highly expressed in Gujing leaves compared with Shuchazao and Siqiu leaves, which may lead to the high accumulation of galloylquinic acid in Gujing leaves. A 14-bp deletion spanning the −765–(−7 5 1) range in the F3′5′H promoter potentially led to low F3′5′H expression levels in Siqiu and Gujing tea plants, which severely disrupted the accumulation of trihydroxy flavonoids in Gujing and Siqiu tea leaves. The high astringency intensity in Gujing tea could be due to the high accumulation of proanthocyanidins and galloylquinic acid. The results of the present study may improve our understanding of the metabolic characteristics of each evolutionary group of species or varieties in the section Thea of the genus Camellia.