Metabolomic and transcriptomice analyses of flavonoid biosynthesis in apricot fruits.

Introduction: Flavonoids, as secondary metabolites in plants, play important roles in many biological processes and responses to environmental factors. Methods: Apricot fruits are rich in flavonoid compounds, and in this study, we performed a combined metabolomic and transcriptomic analysis of orange flesh (JN) and white flesh (ZS) apricot fruits. Results and discussion: A total of 222 differentially accumulated flavonoids (DAFs) and 15855 differentially expressed genes (DEGs) involved in flavonoid biosynthesis were identified. The biosynthesis of flavonoids in apricot fruit may be regulated by 17 enzyme-encoding genes, namely PAL (2), 4CL (9), C4H (1), HCT (15), C3'H (4), CHS (2), CHI (3), F3H (1), F3'H (CYP75B1) (2), F3'5'H (4), DFR (4), LAR (1), FLS (3), ANS (9), ANR (2), UGT79B1 (6) and CYP81E (2). A structural gene-transcription factor (TF) correlation analysis yielded 3 TFs (2 bHLH, 1 MYB) highly correlated with 2 structural genes. In addition, we obtained 26 candidate genes involved in the biosynthesis of 8 differentially accumulated flavonoids metabolites in ZS by weighted gene coexpression network analysis. The candidate genes and transcription factors identified in this study will provide a highly valuable molecular basis for the in-depth study of flavonoid biosynthesis in apricot fruits.

Vitamin B6 as a novel risk biomarker of fractured ankles.

ABSTRACT: Ankle fractures are the most common intra-articular fractures. Osteoporosis is a common and frequent disease among the elderly with a poor prognosis and high risk of fractured ankles. However, the relationship between vitamin B6 and the incidence of fractured ankles in patients with osteoporosis is unclear.A total of 101 patients with osteoporosis were recruited. Clinical and followed-up information was recorded. And the vitamin B6, albumin, globulin, and hemoglobin in the blood were tested. Pearson's chi-squared and spearman test were performed to analyze the correlation between fractured ankles and relative parameters. Univariate and multivariate logistic regression, receiver operating characteristic curve analysis, univariate and multivariate Cox proportional hazards regression analysis, and Kaplan-Meier method were also performed.There exist strong relation between the expression level of vitamin B6 and fractured ankle (P < .001). The expression of vitamin B6 [Odd ratio (OR) = 12.071, 95% confidence interval (CI): 4.69-31.143, P < .001] has a clear correlation with whether the patients have fractured ankles via the univariate logistic regression analysis. In terms of multivariate logistic regression level, vitamin B6 (OR = 15.384, 95% CI:5.195-45.556, P < .001) was significantly associated with fractured ankle. In addition, expression level of vitamin B6 [hazard ratio (HR) = 11.684, 95% CI: 6.419-21.267, P < .001] was significantly associated with Maintenance time from recovery to recurrence (MRTT) of patients with osteoporosis.Enhanced vitamin B6 is significantly correlated with the poor prognosis of patients with osteoporosis and the increasing incidence of fractured ankles.

Identification of Key Genes Controlling Carotenoid Metabolism during Apricot Fruit Development by Integrating Metabolic Phenotypes and Gene Expression Profiles.

To explore the metabolic basis of carotenoid accumulation in different developmental periods of apricot fruits, targeted metabonomic and transcriptomic analyses were conducted in four developmental periods (S1-S4) in two cultivars (Prunus armeniaca cv. "Kuchebaixing" with white flesh and P. armeniaca cv. "Shushangganxing" with orange flesh) with different carotenoid contents. 14 types of carotenes and 27 types of carotene lipids were identified in apricot flesh in different developmental periods. In S3 and S4, the carotenoid contents of the two cultivars were significantly different, and ß-carotene and (E/Z)-phytoene were the key metabolites that caused the difference in the total carotenoid content between the examined cultivars. Twenty-five structural genes (including genes in the methylerythritol 4-phosphate and carotenoid biosynthesis pathways) related to carotenoid biosynthesis were identified among the differentially expressed genes in different developmental periods of the two cultivars, and a carotenoid metabolic pathway map of apricot fruits was drawn according to the KEGG pathway map. The combined analysis of carotenoid metabolism data and transcriptome data showed that PSY, NCED1, and CCD4 were the key genes leading to the great differences in the total carotenoid content. The results provide a new approach to study the synthesis and accumulation of carotenoids in apricot fruits.

Transcriptome analysis insight into ethylene metabolism and pectinase activity of apricot (Prunus armeniaca L.) development and ripening.

Ethylene metabolism is very important for climacteric fruit, and apricots are typical climacteric fruit. The activity of pectinase is closely related to fruit firmness, which further affects fruit quality. To better understand ethylene metabolism, pectinase activity and their molecular regulation mechanisms during the development and ripening of apricot fruit, ethylene metabolism, pectinase activity and the "Luntaibaixing" apricot fruit transcriptome were analyzed at different developmental stages. Ethylene metabolic precursors, enzyme activities and ethylene release increased during fruit development and ripening, with significant differences between the ripening stage and other stages (P < 0.05). Fruit firmness decreased significantly from the S1 to S5 stages, and polygalacturonase, pectin methylesterase, and pectin lyase activities were significantly higher in the S5 stage than in other stages. RNA sequencing (RNA-seq) analysis of fruit resulted in the identification of 22,337 unigenes and 6629 differentially expressed genes (DEGs) during development and ripening, of which 20,989 unigenes are annotated in public protein databases. In functional enrichment analysis, DEGs among the three stages were found to be involved in plant hormone signal transduction. Four key genes affecting ethylene metabolism, six key ethylene signal transduction genes and seven genes related to pectinase in apricot fruit were identified by KEGG pathway analysis. By RNA-sequencing, we not only clarified the molecular mechanism of ethylene metabolism during the ripening of "Luntaibaixing" apricot fruit but also provided a theoretical basis for understanding pectin metabolism in apricot fruit.

Application of Robot Positioning for Cannulated Screw Internal Fixation in the Treatment of Femoral Neck Fracture: Retrospective Study.

BACKGROUND: Femoral neck fracture is a common type of hip fracture. Conventional surgical treatment aims at fixing the fracture site with screws and then gradually promoting bone healing. A robot-assisted orthopedic surgery system is computer technology applied to surgical treatment. OBJECTIVE: This study aimed to explore the therapeutic effect and prognostic value of percutaneous cannulated screw internal fixation using robot-assisted positioning in patients with femoral neck fractures. METHODS: From July 2018 to September 2019, 42 cases of femoral neck fracture admitted to the Second Affiliated Hospital of Luohe Medical College were randomly and averagely divided into control and study groups. The patients in the control group were treated with conventional percutaneous cannulated screw internal fixation, while the patients in the study group were treated with robot-assisted percutaneous cannulated screw fixation during surgical treatment. We compared the treatment conditions and results of the operation between the 2 groups. The Harris score was used to evaluate the treatment efficacy. The state of fracture healing was followed up and compared between the 2 groups. RESULTS: The duration of the operation was shorter, there was less fluoroscopy use, and there were fewer drilled holes in the study group than in the control group (all, P<.001). There was no statistical difference in the amount of intraoperative bleeding between the 2 groups (P=.33). The Harris score (P=.045) and number of excellent and good ratings (P=.01) were significantly higher in the study group than in the control group. The difference in the fracture healing rate between the 2 groups was not statistically significant (P=.23). The fracture healing duration of the study group was shorter than that of the control group (P=.001). CONCLUSIONS: The use of robotic positioning aids in the treatment of femoral neck fractures with percutaneous cannulated screw fixation can effectively improve the efficiency of surgery, shorten the duration of surgery, and reduce the radiation damage to patients. Meanwhile, it improves postoperative treatment and recovery rates of the patients and shortens the fracture healing time.

Analysis of carotenoid content and diversity in apricots (Prunus armeniaca L.) grown in China.

The carotenoids in the peel and flesh of 41 apricot cultivars were qualitatively and quantitatively analysed by UHPLC-APCI-MS/MS, and the L*, a*, b* and quality indexes of the fruits were determined. The results showed that the L*, a*, b* and quality indexes of the fruits were quite different, and 13 carotenoids were detected in the peel and flesh of apricots, among which ε-carotene, α-cryptoxanthin and apocarotenal were newly detected carotenoids in apricots. The total carotenoid content of the 41 apricot cultivars varied from 20.983 to 320.278 µg/g FW, and the total carotenoid content varied from 17.353 to 222.098 µg/g FW in the peel and from 2.536 to 98.179 µg/g FW in the flesh. The main components of apricot fruits were ß-carotene and (E/Z)-phytoene, followed by ß-cryptoxanthin and lutein. This study shows that carotenoids in apricot fruits have rich metabolic diversity.

Genetic diversity, population structure, and relationships of apricot (Prunus) based on restriction site-associated DNA sequencing.

Single-nucleotide polymorphisms (SNPs) are the most abundant form of genomic polymorphisms and are widely used in population genetics research. Here, high-throughput sequencing was used to examine the genome-level diversity, population structure, and relationships of apricot, which are important for germplasm conservation and molecular breeding. Restriction site-associated DNA sequencing (RAD-seq) was adopted to sequence 168 Prunus spp. accessions distributed in five ecological groups, including 74 accessions of cultivated Prunus armeniaca L. and 94 accessions of wild apricots (P. armeniaca L. and Prunus sibirica L.), which generated 417,961 high-quality SNPs. We used cluster, genetic structure, and principal component analyses to examine the genetic diversities and genetic relationships of the 168 accessions. The Dzhungar-Ili ecological group accessions showed the highest genetic diversity in terms of private allele number, observed heterozygosity, and nucleotide diversity. We speculate that the Central Asian ecological group accessions were domesticated from the Dzhungar-Ili ecological group accessions. The population structure and gene flow of the North China and European ecological group accessions suggested a genetic background of P. sibirica. We argue that the two groups should be considered hybrid swarms connected to P. sibirica by continuous and extensive gene flow. P. armeniaca originated in Northwest China (Ili Valley), subsequently spread throughout Central Asia, and eventually spread to Europe. In addition, selective sweep signatures in P. armeniaca during domestication from wild to cultivated apricots, combined with differentially expressed genes, underlie distinct fruit traits, including sugars, aromas, organic acids, and carotenoids. This study provides substantive and valuable genomic resources that will significantly advance apricot improvement and effective utilization.

Identification of key genes and regulators associated with carotenoid metabolism in apricot (Prunus armeniaca) fruit using weighted gene coexpression network analysis.

BACKGROUND: Carotenoids are a class of terpenoid pigments that contribute to the color and nutritional value of many fruits. Their biosynthetic pathways have been well established in a number of plant species; however, many details of the regulatory mechanism controlling carotenoid metabolism remain to be elucidated. Apricot is one of the most carotenoid-rich fruits, making it a valuable system for investigating carotenoid metabolism. The purpose of this study was to identify key genes and regulators associated with carotenoid metabolism in apricot fruit based on transcriptome sequencing. RESULTS: During fruit ripening in the apricot cultivar 'Luntaixiaobaixing' (LT), the total carotenoid content of the fruit decreased significantly, as did the levels of the carotenoids ß-carotene, lutein and violaxanthin (p < 0.01). RNA sequencing (RNA-Seq) analysis of the fruit resulted in the identification of 44,754 unigenes and 6916 differentially expressed genes (DEGs) during ripening. Among these genes, 33,498 unigenes were annotated using public protein databases. Weighted gene coexpression network analysis (WGCNA) showed that two of the 13 identified modules ('blue' and 'turquoise') were highly correlated with carotenoid metabolism, and 33 structural genes from the carotenoid biosynthetic pathway were identified. Network visualization revealed 35 intramodular hub genes that putatively control carotenoid metabolism. The expression levels of these candidate genes were determined by quantitative real-time PCR analysis, which showed ripening-associated carotenoid accumulation. This analysis revealed that a range of genes (NCED1, CCD1/4, PIF3/4, HY5, ERF003/5/12, RAP2-12, AP2, AP2-like, BZR1, MADS14, NAC2/25, MYB1R1/44, GLK1/2 and WRKY6/31/69) potentially affect apricot carotenoid metabolism during ripening. Based on deciphering the molecular mechanism involved in ripening, a network model of carotenoid metabolism in apricot fruit was proposed. CONCLUSIONS: Overall, our work provides new insights into the carotenoid metabolism of apricot and other species, which will facilitate future apricot functional studies and quality breeding through molecular design.

The R2R3-MYB transcription factor PaMYB10 is involved in anthocyanin biosynthesis in apricots and determines red blushed skin.

BACKGROUND: The majority of apricot (Prunus armeniaca L.) cultivars display orange or yellow background skin, whereas some cultivars are particularly preferred by consumers because of their red blushed skin on the background. RESULTS: In this study, two blushed ('Jianali' and 'Hongyu') and two nonblushed ('Baixing' and 'Luntaixiaobaixing') cultivars were used to investigate the formation mechanism of blushed skin in apricots. High-performance liquid chromatography (HPLC) analysis showed that the blushed cultivars accumulated higher cyanidin-3-O-glucoside, cyanidin-3-O-rutinoside and peonidin-3-O-rutinoside levels during fruit ripening than the nonblushed cultivars. Based on coexpression network analysis (WGCNA), a putative anthocyanin-related R2R3-MYB, PaMYB10, and seven structural genes were identified from transcriptome data. The phylogenetic analysis indicated that PaMYB10 clustered in the anthocyanin-related MYB clade. Sequence alignments revealed that PaMYB10 contained a bHLH-interaction motif ([DE]Lx2[RK]x3Lx6Lx3R) and an ANDV motif. Subcellular localization analysis showed that PaMYB10 was a nuclear protein. Real-time qRT-PCR analysis demonstrated that the transcript levels of PaMYB10 and seven genes responsible for anthocyanin synthesis were significantly higher in blushed than in nonblushed apricots, which was consistent with the accumulation of anthocyanin. In addition, bagging significantly inhibited the transcript levels of PaMYB10 and the structural genes in 'Jianali' and blocked the red coloration and anthocyanin accumulation. Transient PaMYB10 overexpression in 'Luntaixiaobaixing' fruits resulted in the red blushed skin at the maturation stage. CONCLUSIONS: Taken together, these data reveal that three anthocyanins are responsible for the blushed skin of apricots, identify PaMYB10 as a positive regulator of anthocyanin biosynthesis in apricots, and demonstrate that blush formation depends on light.

Targeting PRMT5/Akt signalling axis prevents human lung cancer cell growth.

The emerging evidence reveals that protein arginine methyltransferase 5 (PRMT5) is involved in regulation of tumour cell proliferation and cancer development. Nevertheless, the exact role of PRMT5 in human lung cancer cell proliferation and the underlying molecular mechanism remains largely obscure. Here, we showed that PRMT5 was highly expressed in human lung cancer cells and lung cancer tissues. Furthermore, we generated PRMT5 stable knockdown cell lines (A549 and H1299 cells) and explored the functions of PRMT5 in lung cancer cell proliferation. We found that the down-regulation of PRMT5 by shRNA or the inhibition of PRMT5 by specific inhibitor GSK591 dramatically suppressed cyclin E1 and cyclin D1 expression and cell proliferation. Moreover, we uncovered that PRMT5 promoted lung cancer cell proliferation via regulation of Akt activation. PRMT5 was directly co-localized and interacted with Akt, but not PTEN and mTOR. Down-regulation or inhibition of PRMT5 markedly reduced Akt phosphorylation at Thr308 and Ser473, whereas the expression of PTEN and mTOR phosphorylation was unchanged, indicating that PRMT5 was an important upstream regulator of Akt and induced lung cancer cell proliferation. Altogether, our results indicate that PRMT5 promotes human lung cancer cell proliferation through direct interaction with Akt and regulation of Akt activity. Our findings also suggest that targeting PRMT5 may have therapeutic potential for treatment of human lung cancer.

Inhalation lung injury induced by smoke bombs in children: CT manifestations, dynamic evolution features and quantitative analysis.

BACKGROUND: This retrospective study aimed to investigate the computed tomography (CT) manifestations, short-term dynamic evolution features and quantitative lung CT analysis of inhalation lung injury induced by smoke bomb flare. METHODS: Eleven pediatric patients (aged 11 to 13) who inhaled the smoke of smoke bombs underwent several low-dose chest CT scans. The image characteristics and their dynamic changes were observed and quantitative CT values were analyzed. The quantitative CT indicators included lung injury CT score (LICTS), lung fibrosis CT score (LFCTS), mean lung density (MLD), normally aerated volume ratio (NAVR) and reductively aerated volume ratio (RAVR). Box-plot was used to analyze the dynamic changes of each indicator and Spearman statistical method was used to analyze the correlation between any two indicators. RESULTS: (I) In most cases, there were multiple consolidation and massive ground-glass opacities (GGOs) in the two lungs, which aggravated in the early stage and then gradually dissoluted in the later stage. LICTS was positively correlated with MLD (r=0.811, P=0.000), while it was negatively correlated with NAVR (r=-0.712, P=0.000). There existed interstitial fibrosis in the later stage, and LFCTS was positively correlated with RAVR (r=0.382, P=0.028). (II) In one case, the patterns were like layered cake, i.e., consolidation with air bronchus signs in the accumulation area, GGOs in the aforementioned area and normal lung in the top area. The patterns aggravated in the early stage and quickly dissolved in the later stage, and only a few residual fibrotic lesions existed on the final scan. (III) For severe cases, pneumomediastinum and subcutaneous emphysema aggravated in the early stage and then gradually dissolved in the later stage. CONCLUSIONS: The chest CT manifestations of inhalation lung injury induced by smoke bombs are predominantly GGOs and consolidation. They aggravate in the early stage and gradual dissolute in the later stage. CT quantitative values can contribute to evaluating the extent of this disease, and NAVR and RAVR can be used to assess pulmonary function.

[Effects of curcumin on pulmonary fibrosis and functions of paraquat-challenged rats].

OBJECTIVE: To observe the effects of curcumin on pulmonary fibrosis and functions on paraquat (PQ)-challenged rats, and investigate the possible mechanism. METHODS: 108 SPF Wistar rats were divided into three groups according to random number sheet: normal saline (NS) control group, PQ model group and curcumin-treatment group. The rats in each group were subdivided into three subgroups according to different time points (3, 7, 14 days), with 12 rats in each subgroup. PQ-challenged models were reproduced by intragastrical administration of PQ solution 50 mg/kg, and those in NS control group were given the equal volume of NS. After 30 minutes, the rats in curcumin-treatment group were given 200 mg/kg of curcumin by intraperitoneal injection, and those in NS control group and PQ model group were given the equal volume of NS. At 3, 7, 14 days, the tidal volume (VT) was examined, and the blood was drawn from femoral artery for blood gas analysis. Then the rats were sacrificed and the lung tissues were harvested, the hydroxyproline (Hyp) content was measured by alkaline hydrolysis; the expression of transforming growth factor-ß1 (TGF-ß1) was determined by immuno-histochemistry; the distribution and the change of the pulmonary collagen fiber were observed after Masson staining. RESULTS: After exposure to PQ, the VT and arterial partial pressure of oxygen (PaO2) were decreased gradually, and the levels of Hyp and TGF-ß1 were increased gradually, reaching the trough or the peak at 14 days, which were significantly lower or higher than those in NS control group [14-day VT (mL): 1.52±0.33 vs. 2.81±0.47, 14-day PaO2 (kPa): 5.87±0.95 vs. 14.15±1.02, 14-day Hyp (µg/mg): 3.12±0.06 vs. 1.14±0.05, 14-day TGF-ß1 (integral A value): 29.72±4.27 vs. 4.15±0.52, all P < 0.01]. After intervene of curcumin, the parameters were significantly improved as compared with those of PQ model group [14-day VT (mL): 2.34±0.19 vs. 1.52±0.33, 14-day PaO2 (kPa): 10.23±1.01 vs. 5.87±0.95, 14-day Hyp (µg/mg): 2.31±0.04 vs. 3.12±0.06, 14-day TGF-ß1 (integral A value): 15.46±2.89 vs. 29.72±4.27, all P < 0.01]. It was shown by Masson staining that in PQ model group, with the PQ-poisoned time prolonging, diffused pulmonary fibrosis and a large number of collagen deposition were observed gradually, and the most serious collagen deposition was observed at 14 days; after intervene of curcumin, pulmonary fibrosis was alleviated significantly at different time points as compared with the PQ model group. CONCLUSIONS: Curcumin can enhance the pulmonary function by reducing the deposition of collagen fiber and inhabiting pulmonary fibrosis of PQ-poisoned rats.

The effect of siRNA-mediated lymphocyte-specific protein tyrosine kinase (Lck) inhibition on pulmonary inflammation in a mouse model of asthma.

OBJECTIVE: To explore the effect of siRNA-mediated inhibition of lymphocyte-specific protein tyrosine kinase (Lck) on pulmonary inflammation in a mouse model of asthma. METHODS: A total of 32 female BABL/c mice were used in the study. The mouse asthma model was established with ovabumin (OVA), and Lck specific siRNA or nonspecific siRNA was transfected through the tail vein before the first OVA challenge. Two days after the last challenge, mice were sacrificed and bronchoalveolar lavage fluid (BALF), plasma and lung tissue were collected. Levels of Lck mRNA and protein in lung were detected by quantitative real-time PCR and western blot. The levels of IL-4 and IgE in BALF and plasma were detected with ELISA. RESULTS: Lck specific siRNA significantly inhibited expression of Lck mRNA and protein in T cells. In vivo transfection of Lck siRNA down regulated the expression of Lck mRNA and protein in lung parenchymal homogenates. Sensitized mice treated with Lck siRNA prior to OVA challenge had fewer eosinophils in BALF and in lung sections and lower levels of IL-4 and IgE in BALF and plasma compared to those treated with nonspecific siRNA. CONCLUSIONS: Pretreatment of OVA sensitized mice with Lck siRNA results in attenuation of pulmonary inflammation following OVA challenge. Inhibition of Lck gene expression should be investigated further as a potential therapy for asthma.

[Fragmentation process of wetland landscape in watersheds of Sanjiang Plain, China].

The Sanjiang Plain is the largest fresh water wetland distribution area in China and the center of waterfowls breeding and habitat area in Asia, but over the past 50 years, more than 73% of its wetland had lost because of agricultural development, and as a result, the wetland biodiversity declines dramatically, and the remnant wetlands are in a very fragment state. Based on historical maps, remote sensing data and GIS techniques, this paper selected two watersheds to analyze their wetland landscape fragmentation process during 1950-2000. It was indicated that land reclamation resulted in a decrease of 98% wetland corridors in Qixing River, 90% in Naoli River, 87% in the middle reach of Bielahong River, and 94% in the lower reach of Bielahong River; The amount of isolated wetlands in watershed increased dramatically; The maximum patch areas of wetland decreased by 92.6% in Naoli River watershed and 74.6% in Bielahong River watershed, and the mean wetland patch area in the two watersheds decreased by 99%. Before 1983, the wetland landscape was in an extensive area distribution state (the index of patch density was < 0.1), but after 1983, it fragmented dramatically, with the index of patch density larger than 1.5. The shape fragmentation indices of wetland decreased from 1950 to 2000, indicating a very big change in wetland patch shapes in the watersheds. The area fragmentation indices of wetland also increased from 1950 to 2000, especially after 1983, showing that the wetlands were in a serious fragmentation state. The wetland landscape fragmentation changed from a landmass and island model to a satellite model, and finally to a completely isolated model, which indicated the great changes in spatial structure of wetland in the Sanjiang Plain.

Impacts on wetlands of large-scale land-use changes by agricultural development: the Small Sanjiang Plain, China.

The Small Sanjiang Plain (SSP), was formerly the largest wetland complex in China, located in the Northeastern part of Heilongjiang Province, China. Home to vast numbers of waterfowls, fish, and plants, the SSP is globally significant for biodiversity conservation. The loss and fragmentation of wetlands as a result agricultural development over 50 years has impacted wetland communities and their biodiversity. We used GIS to inventory large-scale land-use changes from 1950 to 2000, together with other statistical data. We found that 73.6% of the wetlands were lost due to agricultural development. Consequences of these land-use changes included: i) a rapid decline in waterfowl and plant species with the loss and fragmentation of natural wetlands and wetland ecosystem degradation; ii) greater variation in wetland water levels as the result of land-use changes over the years; iii) disruption of the dynamic river-floodplain connection by construction of drainage ditches and levees; and iv) a decrease in floodplain area that caused increased flooding peak flows and runoff. Here we show how these changes affect wetland biodiversity and impact important wetland species.