STAY-GREEN Accelerates Chlorophyll Degradation in Magnolia sinostellata under the Condition of Light Deficiency.

Species of the Magnoliaceae family are valued for their ornamental qualities and are widely used in landscaping worldwide. However, many of these species are endangered in their natural environments, often due to being overshadowed by overstory canopies. The molecular mechanisms of Magnolia's sensitivity to shade have remained hitherto obscure. Our study sheds light on this conundrum by identifying critical genes involved in governing the plant's response to a light deficiency (LD) environment. In response to LD stress, Magnolia sinostellata leaves were endowed with a drastic dwindling in chlorophyll content, which was concomitant to the downregulation of the chlorophyll biosynthesis pathway and upregulation in the chlorophyll degradation pathway. The STAY-GREEN (MsSGR) gene was one of the most up-regulated genes, which was specifically localized in chloroplasts, and its overexpression in Arabidopsis and tobacco accelerated chlorophyll degradation. Sequence analysis of the MsSGR promoter revealed that it contains multiple phytohormone-responsive and light-responsive cis-acting elements and was activated by LD stress. A yeast two-hybrid analysis resulted in the identification of 24 proteins that putatively interact with MsSGR, among which eight were chloroplast-localized proteins that were significantly responsive to LD. Our findings demonstrate that light deficiency increases the expression of MsSGR, which in turn regulates chlorophyll degradation and interacts with multiple proteins to form a molecular cascade. Overall, our work has uncovered the mechanism by which MsSGR mediates chlorophyll degradation under LD stress conditions, providing insight into the molecular interactions network of MsSGR and contributing to a theoretical framework for understanding the endangerment of wild Magnoliaceae species.

Metabolic analysis of the regulatory mechanism of sugars on secondary flowering in Magnolia.

BACKGROUND: Magnolia, a traditional and important ornamental plant in urban greening, has been cultivated for about 2000 years in China for its elegant flower shape and gorgeous flower color. Most varieties of Magnolia bloom once a year in spring, whereas a few others, such as Magnolia liliiflora Desr. 'Hongyuanbao', also bloom for the second time in summer or early autumn. Such a twice flowering trait is desirable for its high ornamental value, while its underlying mechanism remains unclear. METHODS: Paraffin section was used to show the flowering time and phenotypic changes of M. liliiflora 'Hongyuanbao' during the twice flowering periods from March 28 to August 25, 2018. Gas chromatography-mass spectrometry (GC-MS) was then performed to explore the chemical metabolites through the twice flower bud differentiation process in 'Hongyuanbao', and the metabolites were screened and identified by orthogonal projection to latent structures discriminant analysis (OPLS-DA). Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis (KEGG) was used to reveal the relationship between the sugar metabolites and twice-flowering characteristic. To further investigate the potential role of sucrose and trehalose on flowering regulation of 'Hongyuanbao', the plants once finished the spring flowering were regularly sprayed with sucrose and trehalose solutions at 30 mM, 60 mM, and 90 mM concentrations from April 22, 2019. The flower bud differentiation processes of sprayed plants were observed and the expression patterns of the genes involved in sucrose and trehalose metabolic pathways were studied by quantitative reverse transcription PCR (qRT-PCR). RESULTS: It showed that 'Hongyuanbao' could complete flower bud differentiation twice in a year and flowered in both spring and summer. The metabolites of flower bud differentiation had a significant variation between the first and second flower buds. Compared to the first flower bud differentiation process, the metabolites in the sucrose and trehalose metabolic pathways were significantly up-regulated during the second flower bud differentiation process. Besides that, the expression levels of a number of trehalose-6-phosphate synthase (TPS) genes including MlTPS1, MlTPS5, MlTPS6, MlTPS7 and MlTPS9 were substantially increased in the second flower differentiation process compared with the first process. Exogenous treatments indicated that compared to the control plants (sprayed with water, CK), all three concentrations of trehalose could accelerate flowering and the effect of 60 mM concentration was the most significant. For the sucrose foliar spray, only the 60 mM concentration accelerated flowering compared with CK. It suggested that different concentration of trehalose and sucrose might have different effects. Expression analysis showed that sucrose treatment increased the transcription levels of MlTPS5 and MlTPS6, whereas trehalose treatment increased MlTPS1, showing that different MlTPS genes took part in sucrose and trehalose metabolic pathways respectively. The expression levels of a number of flowering-related genes, such as MlFT, MlLFY, and MlSPL were also increased in response to the sprays of sucrose and trehalose. CONCLUSIONS: We provide a novel insight into the effect of sucrose and trehalose on the flowering process in Magnolia. Under the different sugar contents treatments, the time of flower bud differentiation of Magnolia was advanced. Induced and accelerated flowering in response to sucrose and trehalose foliar spray, coupled with elevated expression of trehalose regulatory and response genes, suggests that secondary flower bud formation is a promoted by altered endogenous sucrose and trehalose levels. Those results give a new understanding of sucrose and trehalose on twice-flowering in Magnolia and provide a preliminary speculation for inducing and accelerating the flowering process in Magnolia.

Identification of potential light deficiency response regulators in endangered species Magnolia sinostellata.

Magnolia sinostellata is one of the endangered species in China and largely suffers light deficiency stress in the understory of forest. However, the weak light response molecular mechanism remains unclear. More importantly, hub genes in the molecular network have not been pinpointed. To explore potential regulators in the mechanism, weighted gene co-expression network analysis (WGCNA) was performed to analysis the trancriptome data of M. sinostellata leaves subjected to weak light with different time points. Gene co-expression analysis illustrated that module 1, 2 and 3 were closely associated with light deficiency treatment, which. Gene ontology and KEGG analyses showed that genes in module 1 mainly participated in amino and nucleotide metabolism, module 2 mostly involved in carbon fixation and module 3 mostly regulated photosynthesis related pathways, among which 6, 7 and 8 hub genes were identified, respectively. Hub genes isoform_107196 in module 1 and isoform_55976 in module 2 were unique to M. sinostellata. This study found that light deficiency inhibited photosynthesis and stress tolerance, while improved carbon metabolism and flowering related pathways in M. sinostellata, which can impact its accumulation reserves of growth and reproduction in the next season. In addition, key shade response regulators identified in this study have laid a firm foundation for further investigation of shade response molecular mechanism and protection of other shade sensitive plants.

A nomogram model predicting the risk of postpartum stress urinary incontinence in primiparas: A multicenter study.

Stress urinary incontinence (SUI) is a common gynecological urinary system disease, and globally, 200 million or more people suffer from it. However, the existing literature mostly focuses on postpartum urinary incontinence (UI) or UI in middle-aged and elderly people, with little focus on primiparas. To analyse urinary incontinence prevalence and its risk factors in primiparas and establish a nomogram prediction model, 360 parturients were recruited from three hospitals between April and September 2021. A homemade electronic questionnaire was used to investigate the general demographic and perinatal characteristics of primiparas. The SUI was diagnosed by the physicians. Logistic regression analysis of independent risk factors for SUI and a nomogram prediction model were established. Ninety people were diagnosed as SUI. The number of pregnancies (OR = 3.322, 95% CI = 1.473-7.492), residence (OR = 5.451, 95% CI = 2.725-10.903), occupation (OR = 3.393, 95% CI = 1.144-10.064), education level (OR = 3.551, 95% CI = 1.223-10.308), delivery method (OR = 10.270, 95% CI = 4.090-25.789), and oxytocin use (OR = 2.166, 95% CI = 1.142-4.109) were independent risk factors for SUI. The C-index of the nomogram prediction model was 0.798 (95% CI = 0.749-0.846). The POPDI score, CRADI score, UDI score, and PFDI scores of women with SUI were significantly higher than those of non-SUI women, while I-QOL scores were significantly lower than those of non-SUI women. In conclusion, the prevalence of SUI among primiparas in Fuyang, China, was 25.00%, which exhibited a large impact on the quality of life of puerperae. The present study successfully established an individualized nomogram prediction model of SUI for primiparas with good discrimination and diagnostic efficiency, which was helpful for the early clinical identification of high-risk primiparas with SUI.

Metachronous Multiple Primary Carcinoma With Acute Promyelocytic Leukemia: 2 Cases Report and Literature Review.

The co-occurrence of multiple primary cancers with hematological malignancies is uncommon, and acute promyelocytic leukemia (APL) with MPC is even rarer, with only a few cases reported in the literature. Herein, we introduce the diagnosis and treatment of 2 cases of MPC complicated with APL in our hospital and review the relevant literature. Both patients were primary solid tumor patients and were treated with surgery and chemotherapy, and had stable disease (SD). However, more than 1 year after the primary tumor was diagnosed, clinical symptoms were found and APL was diagnosed. Both patients received standard remission-induction therapy, but unfortunately died in the short term due to hemorrhagic complications. In conclusion, treatment of hematological neoplasms, especially acute leukemia combined with multiple primary cancers, is challenging. The prognostic factors and survival analysis of MPC patients with combined APL still need further clinical research and analysis.

Irisin mediates beiging of adipose-derived mesenchymal stem cells through binding to TRPC3.

BACKGROUND: Beiging of white fat plays an important role in energy metabolism. Beige adipocytes contribute to the regulation of body weight and body temperature through expenditure of chemical energy to produce heat, and they have therefore recently attracted considerable attention as potential targets for therapeutic approaches in metabolic disorders, including obesity. All adipocytes, including beige adipocytes, differentiate from mesenchymal stem cells (MSCs), which may provide an important path for clinical intervention; however, the mechanism of beiging of human adipose cell-derived MSCs is not fully understood. Here, we provide insights on the role of IRISIN, which is known to be secreted by skeletal muscle and promote beiging of white fat. RESULTS: We established an IRISIN-induced mesenchymal stem cell beiging model and found that IRISIN protein interacts with the MSC membrane protein TRPC3. This interaction results in calcium influx and consequential activation of Erk and Akt signaling pathways, which causes phosphorylation of PPARγ. The phosphorylated PPARγ enters the nucleus and binds the UCP1 promoter region. Furthermore, the role of TRPC3 in the beiging of MSCs was largely abolished in Trpc3-/- mice. We additionally demonstrate that the calcium concentration in the brain of mice increases upon IRISIN stimulation, followed by an increase in the content of excitatory amino acids and norepinephrine, while Trpc3-/- mice exhibit the reverse effect. CONCLUSIONS: We found that TRPC3 is a key factor in irisin-induced beiging of MSCs, which may provide a new target pathway in addressing metabolic disorders. Our results additionally suggest that the interaction of irisin with TRPC3 may affect multiple tissues, including the brain.

Light Deficiency Inhibits Growth by Affecting Photosynthesis Efficiency as well as JA and Ethylene Signaling in Endangered Plant Magnolia sinostellata.

The endangered plant Magnolia sinostellata largely grows in the understory of forest and suffers light deficiency stress. It is generally recognized that the interaction between plant development and growth environment is intricate; however, the underlying molecular regulatory pathways by which light deficiency induced growth inhibition remain obscure. To understand the physiological and molecular mechanisms of plant response to shading caused light deficiency, we performed photosynthesis efficiency analysis and comparative transcriptome analysis in M. sinostellata leaves, which were subjected to shading treatments of different durations. Most of the parameters relevant to the photosynthesis systems were altered as the result of light deficiency treatment, which was also confirmed by the transcriptome analysis. Gene Ontology and KEGG pathway enrichment analyses illustrated that most of differential expression genes (DEGs) were enriched in photosynthesis-related pathways. Light deficiency may have accelerated leaf abscission by impacting the photosynthesis efficiency and hormone signaling. Further, shading could repress the expression of stress responsive transcription factors and R-genes, which confer disease resistance. This study provides valuable insight into light deficiency-induced molecular regulatory pathways in M. sinostellata and offers a theoretical basis for conservation and cultivation improvements of Magnolia and other endangered woody plants.

Cell-Seeded Acellular Artery for Reconstruction of Long Urethral Defects in a Canine Model.

The management of urethral stricture remains a major therapeutic challenge in clinics. Herein, we explored the feasibility of reconstructing a relatively long segment of the urethra by the cell-seeded acellular artery in a canine model. The acellular arterial matrix was obtained from the excised carotid artery of donor dogs. Autologous adipose-derived stem cells (ADSCs) from 6 male dogs were grown and seeded onto the premade acellular arterial matrix. A 3 cm long segment of the urethra was resected in 12 male dogs. Urethroplasty was performed with the acellular arterial matrix seeded with ADSCs in 6 animals and without cells in 6. Serial urethrography was performed at 1 and 3 months postoperatively. Wide urethral calibers without any signs of strictures were confirmed in all 6 animals in the experimental group. In contrast, urethral stricture was demonstrated in 3 animals in the control group. The graft was highly epithelialized and smooth in the experimental group, while graft contracture and scar formation were showed in the control group. Histologic analysis of the cell-seeded arterial matrix at 1 month confirmed the presence of multilayered urothelium and muscle. The levels of tissue formation developed over time with a progressive increase in muscle content. In contrast, extensive fibrosis and sparse smooth muscle were seen in animals treated with matrix without ADSCs. This study provides preclinical evidence that the ADSC-seeded arterial matrix can be used as a tubularized scaffold in the reconstruction of 3 cm long urethral defect in a male canine model. The ADSC-seeded arterial matrix remodels and regenerates normal-appearing urethral tissue layers over time.

The genome of Magnolia biondii Pamp. provides insights into the evolution of Magnoliales and biosynthesis of terpenoids.

Magnolia biondii Pamp. (Magnoliaceae, magnoliids) is a phylogenetically, economically, and medicinally important ornamental tree species widely grown and cultivated in the north-temperate regions of China. Determining the genome sequence of M. biondii would help resolve the phylogenetic uncertainty of magnoliids and improve the understanding of individual trait evolution within the Magnolia genus. We assembled a chromosome-level reference genome of M. biondii using ~67, ~175, and ~154 Gb of raw DNA sequences generated via Pacific Biosciences single-molecule real-time sequencing, 10X Genomics Chromium, and Hi-C scaffolding strategies, respectively. The final genome assembly was ~2.22 Gb, with a contig N50 value of 269.11 kb and a BUSCO complete gene percentage of 91.90%. Approximately 89.17% of the genome was organized into 19 chromosomes, resulting in a scaffold N50 of 92.86 Mb. The genome contained 47,547 protein-coding genes, accounting for 23.47% of the genome length, whereas 66.48% of the genome length consisted of repetitive elements. We confirmed a WGD event that occurred very close to the time of the split between the Magnoliales and Laurales. Functional enrichment of the Magnolia-specific and expanded gene families highlighted genes involved in the biosynthesis of secondary metabolites, plant-pathogen interactions, and responses to stimuli, which may improve the ecological fitness and biological adaptability of the lineage. Phylogenomic analyses revealed a sister relationship of magnoliids and Chloranthaceae, which are sister to a clade comprising monocots and eudicots. The genome sequence of M. biondii could lead to trait improvement, germplasm conservation, and evolutionary studies on the rapid radiation of early angiosperms.

Low platelet count is a risk factor of postoperative pneumonia in patients with type A acute aortic dissection.

BACKGROUND: Previous studies have already established that low platelet count is related to adverse outcomes in patients with type A acute aortic dissection (AAAD). However, there are yet limited studies investigating the association of platelet count and the risk of postoperative pneumonia in AAAD patients. METHODS: This retrospective cohort study was conducted in Xiangya Hospital of Central South University from January 2014 to May 2019. Clinical and laboratory data were collected. The correlation between platelet count and postoperative pneumonia was analyzed using multivariate logistic regression and the area under the receiver operating characteristic curve (AUC) was used to assess the predictive power of platelet count on pneumonia. RESULTS: A total of 268 patients with AAAD were enrolled. The overall incidence of pneumonia was 36.94% (n=99). Multivariate logistic regression revealed that platelet count was negatively associated with the risk of postoperative pneumonia (OR 0.93; 95% CI: 0.88-0.98) after adjusting for the confounders. Compared to the lowest platelet count tertile (T1), medium platelet count (T2) and highest platelet count (T3) had a lower risk of postoperative pneumonia after adjusting for the confounders (OR 0.80, 95% CI: 0.40-1.60; OR 0.30, 95% CI: 0.13-0.66; respectively). A similar trend was observed when the platelet count was handled as categorical variables (tertiles). The area under the ROC curve was 0.635 (95% CI: 0.565-0.707), with a sensitivity of 76.77%, a specificity of 50.89% and an accuracy of 60.45%. CONCLUSIONS: Our findings indicate that low platelet count is an independent risk factor of postoperative pneumonia in patients with AAAD and has a specific predictive power on the risk of postoperative pneumonia.

A new strategy for accurate targeted diagnosis and treatment of cutaneous malignant melanoma: dual-mode phase-change lipid nanodroplets as ultrasound contrast agents.

BACKGROUND: Currently, effective detection and treatment of cutaneous malignant melanoma (CMM) still face severe challenges. Ultrasound molecular imaging as a noninvasive and easy-to-operate method is expected to bring improvements for tumor detection. PURPOSE: The aim of this research is to prepare novel phase-change ultrasound contrast agents, Nds-IR780, which can perform not only dual-mode molecule-targeted imaging but also targeted photothermal therapy for CMM. METHODS: A double emulsion process was used to prepare the Nds-IR780. Then, the entrapment rate and drug loading of IR-780 iodide in Nds-IR780 were detected by high-performance liquid chromatography. The biocompatibility of Nds-IR780 was evaluated by a CCK-8 assay and the characteristics and stability of that were verified through the particle size analyzer, laser scanning confocal microscopy (LSCM) and transmission electron microscopy (TEM). The abilities of dual-mode molecule-targeted imaging and targeted photothermal therapy for Nds-IR780 were confirmed via the in vitro and in vivo experiments. RESULTS: Nds-IR780 had good size distribution, polydispersity index, stability and biosafety. The in vitro and in vivo experiments confirmed that Nds-IR780 were capable of targeting CMM cells with high affinity (22.4±3.2%) and facilitating dual-mode imaging to detect the primary lesion and sentinel lymph nodes (SLNs) of CMM. Furthermore, the photothermal ablation of CMM mediated by Nds-IR780 was very effective in vivo. CONCLUSION: The newly prepared Nds-IR780 were observed to be effective targeted theranostic probe for the precise detection and targeted treatment of CMM.

FA-NBs-IR780: Novel multifunctional nanobubbles as molecule-targeted ultrasound contrast agents for accurate diagnosis and photothermal therapy of cancer.

Early accurate diagnosis and targeted therapy for cancer are essential to improve the prognosis of patients. With the emergence of molecular imaging, molecule-targeted ultrasound imaging for the non-invasive and precise detection of cancer has attracted increased attention. The investigation of molecule-targeted ultrasound contrast agents (UCAs) with excellent performance is urgently needed. In this study, we synthetized folic acid and IR-780 on self-made nanobubbles and prepared novel UCAs, named FA-NBs-IR780. The results showed that the conjugates had a uniform size distribution (591 ±â€¯52 nm). In vitro and in vivo experiments demonstrated that FA-NBs-IR780 can target tumour cells via dual molecular targeting, perform enhanced-contrast ultrasound imaging and near-infrared fluorescence (NIRF) imaging for the precise detection of tumours, and induce targeted photothermal therapy in lesions irradiated at 808 nm. Ex vivo experiments further confirmed that FA-NBs-IR780 efficiently induced tumour cell apoptosis and inhibited tumour growth. The newly fabricated FA-NBs-IR780 were observed to be molecule-targeted dual-mode UCAs and may have potential applications in early accurate diagnosis and targeted therapy of cancer in the future.

Effects of Gastric Cancer Cell-Derived Exosomes on the Immune Regulation of Mesenchymal Stem Cells by the NF-kB Signaling Pathway.

Mesenchymal stem cells (MSCs) are important components of the tumor microenvironment, which play an important role in tumor development. Exosomes derived from tumor cells can affect the biological characteristics of MSCs. Our study examined the effects of exosomes derived from gastric cancer cells on MSC immunomodulatory functions. Exosomes were extracted from gastric cancer cell line AGS (AGS-Exos) and cultured with MSCs. MSCs were then cocultured with both human peripheral blood mononuclear cells and macrophages [phorbol-12-myristate-13-acetate (PMA)-stimulated THP1 cells]. The activation levels of T cells and macrophages were detected by flow cytometry and real-time quantitative polymerase chain reaction (RT-PCR). Changes in the MSC signaling pathway after AGS-Exos stimulation were studied using RNA Chip, and the molecular mechanisms of functional change in MSCs were studied by inhibiting the signaling pathway. MSCs treated with AGS-Exos could promote macrophage phagocytosis and upregulate the secretion of proinflammatory factor, and promote the activation of CD69 and CD25 on the surface of T cells. RNA Chip results indicated the abnormal activation of the NF-kB signaling pathway in MSCs after AGS-Exos stimulation, and this was verified by the identification of key proteins in the pathway using western blot analysis. After NF-kB signaling pathway inhibition, the effect of MSCs stimulated by AGS-Exos on T cells and macrophages was markedly weakened. Therefore, AGS-Exos affected the immunomodulation function of MSCs through the NF-kB signaling pathway, which enhanced the ability of MSCs to activate immune cells, maintain the inflammatory environment, and support tumor growth.

Development of EST-SSR markers and their application in an analysis of the genetic diversity of the endangered species Magnolia sinostellata.

Magnolia sinostellata is an endemic species of Magnoliaceae that is narrowly distributed in the south of Zhejiang Province, China. To explore the genetic diversity and population structure of this endangered species, this study developed sequence tag-simple sequence repeat (EST-SSR) markers based on transcriptome data of M. sinostellata. In total, 25472 SSRs were identified among 110644 unique assembled sequences with a total of 90.83 Mb and an average frequency of 23.02%. The mononucleotide (33.53%) and dinucleotide (42.08%) motifs appeared to be the most abundant. In total, 150 potential loci were randomly selected to validate the quality of the developed SSR markers; an effective PCR rate of 32.00% and a polymorphism rate of 15.33% were obtained for these loci. After performing sequencing and cloning for validation, 23 pairs of SSR primers were retained and used to characterize the genetic diversity and population structure of M. sinostellata. Overall, 204 alleles were amplified. The results of Shannon's information index (I), heterozygosity (Ho), heterozygosity (He) and Nei's expected heterozygosity (H) indicated rich genetic diversity in M. sinostellata. However, the high inbreeding coefficient and differential coefficient suggest that serious genetic drift occurred within populations, and genetic differentiation is apparent among the populations. Consequently, although M. sinostellata has high genetic diversity among populations, it is still in a serious and dangerous condition. Habitat destruction caused by human activities is the main threat to this species, and enhancing the species abundance by adopting some conservation measures should be favourable for saving the species.

A Novel Bimodal Imaging Agent Targeting HER2 Molecule of Breast Cancer.

Nanobubble (NB), a newly developed nanoscaled ultrasound contrast agent (UCA) for molecular imaging, has been widely researched for these years. Targeting it with functional molecule, nanobubble can adhere selectively to cellular epitopes and receptors outside the vasculature via enhanced permeability and retention (EPR) effect of tumor blood vessel. To enhance the targeting rate of our previous prepared NBs-Affibody for HER2 (+) breast cancer imaging, we introduced a near-infrared fluorescent (NIRF) dye, IR783, in this study to enhance tumor-specific targeting rate and provide a promising modality for dual-mode imaging. The prepared IR783-NBs-Affibody presented a uniform nanoscale size around 482.7 ± 54.3 nm, good biosecurity, and stability over time. The encapsulation efficiency (EE) of IR-783 was 15.09% in the conjugates leading to a successful NIR fluorescence and ultrasound enhancement imaging ex vivo. IR783-NBs-Affibody was able to automatically accumulate on BT474 cells with a highly increased targeting rate of 85.4% compared with previous NBs-Affibody of 26.6%, while Affibody-guided HER2 binding was only found in HER2-positive cell lines (BT474 and T-47D). The newly developed IR783-NBs-Affibody is characterized with favorable HER2 targeting ability and bimodal imaging capability for breast cancer. Thus, IR783-NBs-Affibody holds great potential in molecular diagnosis for patients with breast cancer.

Characterization of new microsatellite markers based on the transcriptome sequencing of Clematis finetiana.

BACKGROUND: Clematis is the biggest genus in the family Ranunculaceae with about 300 species. Clematis is also a globally important commercial group of flowers, especially in the United States and European countries. Their petals with different colors and shapes make the genus the "Queen of the Vines". However, the genomic information and phylogeny of Clematis based on existing molecular studies are limited. In this paper, new microsatellites (SSR) markers were identified from the transcriptome data of C. finetiana obtained using the Illumina paired-end sequencing technology. RESULTS: Sequences on a total of 71,900 high-quality unigenes with the mean length of 865 bp were produced in this study. There were 6192unigenes annotated and classified into 49 functional sub-groups in three main ontology categories in GO (Gen Ontology) database,14,022 unigenes mapped to COGs (Clusters of Orthologous Groups) database and classified into 25 functional categories, and 21,494 unigenes obtained and divided into 128 pathways of KEGG (Kyoto Encyclopedia of Genes) Database. A total of 7532 SSRs were discovered from 6337 unigenes. We randomly tested 210 primer pairs, of which 52 primer pairs were able to generate specific products, and 19 possessed polymorphism in the 13 wild populations of six species from Clematis, which were used as a test material. CONCLUSIONS: The dataset of C. finetiana transcriptome and the identified new SSR markers will promote genetic research and breeding effort in Clematis.

Transcriptomic Analysis of Flower Bud Differentiation in Magnolia sinostellata.

Magnolias are widely cultivated for their beautiful flowers, but despite their popularity, the molecular mechanisms regulating flower bud differentiation have not been elucidated. Here, we used paraffin sections and RNA-seq to study the process of flower bud differentiation in Magnolia sinostellata. Flower bud development occurred between 28 April and 30 May 2017 and was divided into five stages: undifferentiated, early flower bud differentiation, petal primordium differentiation, stamen primordium differentiation, and pistil primordium differentiation. A total of 52,441 expressed genes were identified, of which 11,592 were significantly differentially expressed in the five bud development stages. Of these, 82 genes were involved in the flowering. In addition, MADS-box and AP2 family genes play critical roles in the formation of flower organs and 20 differentially expressed genes associated with flower bud differentiation were identified in M. sinostellata. A qRT-PCR analysis verified that the MADS-box and AP2 family genes were expressed at high levels during flower bud differentiation. Consequently, this study provides a theoretical basis for the genetic regulation of flowering in M. sinostellata, which lays a foundation for further research into flowering genes and may facilitate the development of new cultivars.

Exosomes Derived from Hypoxia-Treated Human Adipose Mesenchymal Stem Cells Enhance Angiogenesis Through the PKA Signaling Pathway.

Angiogenesis is a complicated and sequential process that plays an important role in different physiological processes. Mesenchymal stem cells (MSCs), which are pluripotent stem cells, are widely used for the treatment of ischemic and traumatic diseases, and exosomes derived from these cells can also promote angiogenesis. Therefore, we aimed to uncover mechanisms to improve MSC exosome-mediated angiogenesis. For this study, we isolated human adipose-derived MSCs (hAD-MSCs) and assessed differentiation ability and markers. Cells were divided into hypoxia-treated MSCs (H-MSCs) and normoxia-treated MSCs (N-MSC), and exosomes were extracted by ultrafiltration. Exosomes (100 µg/mL) from H-MSCs and N-MSCs were added to human umbilical vein endothelial cells (HUVECs). Exosome uptake and the ability of endothelial cells to form tubes were detected in real time. Protein samples were collected at different time points to detect the expression of inhibitors (Vash1) and enhancers (Angpt1 and Flk1) of angiogenesis; we also assessed their related signaling pathways. We found that exosomes from the hypoxia group were more easily taken up by HUVECs; furthermore, their angiogenesis stimulatory activity was also significantly enhanced compared to that with exosomes from the normoxia group. HUVECs exposed to exosomes from H-MSCs significantly upregulated angiogenesis-stimulating genes and deregulated angiogenesis-inhibitory genes. The expression of vascular endothelial growth factor (VEGF) and activation of the protein kinase A (PKA) signaling pathway in HUVECs were significantly increased by hypoxia-exposed exosomes. Moreover, a PKA inhibitor was shown to significantly suppress angiogenesis. Finally, we concluded that hypoxia-exposed exosomes derived from hAD-MSCs can improve angiogenesis by activating the PKA signaling pathway and promoting the expression of VEGF. These results could be used to uncover safe and effective treatments for traumatic diseases.