A review of changes at the phenotypic, physiological, biochemical, and molecular levels of plants due to high temperatures.

MAIN CONCLUSION: This review summarizes the physiological, biochemical, and molecular regulatory network changes in plants in response to high temperature. With the continuous rise in temperature, high temperature has become an important issue limiting global plant growth and development, affecting the phenotype and physiological and biochemical processes of plants and seriously restricting crop yield and tree growth speed. As sessile organisms, plants inevitably encounter high temperatures and improve their heat tolerance by activating molecular networks related to heat stress, such as signal transduction, synthesis of metabolites, and gene expression. Heat tolerance is a polygenic trait regulated by a variety of genes, transcription factors, proteins, and metabolites. Therefore, this review summarizes the changes in physiological, biochemical and molecular regulatory networks in plants under high-temperature conditions to lay a foundation for an in-depth understanding of the mechanisms involved in plant heat tolerance responses.

Construction of Frustrated Lewis Pairs in Poly(heptazine imide) Nanosheets via Hydrogen Bonds for Boosting CO2 Photoreduction.

The creation of frustrated Lewis pairs on catalyst surface is an effective strategy for tuning CO2 activation. The critical step in the formation of frustrated Lewis pairs is the spatial effect of proximal Lewis acid-Lewis base pairs. Here, we demonstrate a facile surface functionalization methodology that enables hydrogen bonding between N and H atoms to mediate the construction of frustrated Lewis pairs in poly(heptazine imide), thereby increasing the propensity to activate CO2 molecules. Experimental and theoretical results show that the construction of active hydrogen bonding regions can facilitate the bending of CO2 molecules. Furthermore, the delocalization of electron clouds induced by the hydrogen bonding-mediated frustrated Lewis pairs can promote the heterolytic cleavage and photocatalytic conversion of CO2. This work highlights the potential of utilizing hydrogen bonding-mediated strategy in heterogeneously photocatalytic activation of CO2 over polymer materials.

Pathogenic bi-allelic variants of meiotic ZMM complex gene SPO16 in premature ovarian insufficiency.

Premature ovarian insufficiency (POI) is a heterogeneous disease affecting the physical and mental health of millions of women worldwide. The contribution of genetic factors in the pathogenesis of POI has increased, with quite a few of causative genes involved in meiosis. ZMM proteins are a group of conserved proteins participating in meiotic synapsis and crossover maturation. Here, by screening the variations of ZMM genes in our in-house WES database of 1030 idiopathic POI patients, one novel homozygous variation in SPO16 (c.160 + 8A > G) was firstly identified in one patient. The variation was verified to disturb mRNA splicing by minigene assay, produced a non-functional SPO16 protein, and was classified as pathogenetic according to American College of Medical Genetics guideline. During meiotic prophase I, SHOC1 binds to branched DNA and recruits SPO16 and other ZMM proteins to facilitate crossover formation. Together with our recent identified bi-allelic variations of SHOC1 in a published work, this study highlighted the essential roles of ZMM genes in the maintenance of ovarian function and expanded the POI gene spectrum.

Analysis of the Current Situation and Influencing Factors of Coping Styles in Ischemic Stroke Patients.

Objective: To explore the current situation and influencing factors of coping styles in ischemic stroke patients. Methods: 250 ischemic stroke patients admitted to our hospital from September 2019 to September 2021 were selected as the study subjects to obtain the general information of patients, and the corresponding indexes of patients were evaluated by the method of document investigation to analyze the current situation and influencing factors of coping styles in ischemic stroke patients. Results: Under stressful conditions, patients with no dependence on life-support level, without anxiety and depression, enjoying a high quality of life, and with high self-efficacy were more likely to adopt the positive coping styles (P < .05). Logistic regression analysis showed that infarction area, life-support level, and self-efficacy were independent risk factors for coping style in patients with ischemic stroke (all P < .05). Conclusion: Ischemic stroke patients tend to adopt negative coping styles. Infarction area, life-support level, and self-efficacy of ischemic stroke patients are found to be the main factors affecting their coping styles.

Ellagic Acid from Hull Blackberries: Extraction, Purification, and Potential Anticancer Activity.

Ellagic acid (EA) is present at relatively high concentrations in many berries and has many beneficial health effects, including anticancer properties. To improve the development and utilization of blackberry fruit nutrients, we divided Hull blackberry fruits into five growth periods according to color and determined the EA content in the fruits in each period. The EA content in the green fruit stage was the highest at 5.67 mg/g FW. Single-factor tests and response surface methodology were used to optimize the extraction process, while macroporous resin adsorption and alkali dissolution, acid precipitation, and solvent recrystallization were used for purification. The highest purity of the final EA powder was 90%. The anticancer assessment results determined by MTT assay showed that EA inhibited HeLa cells with an IC50 of 35 µg/mL, and the apoptosis rate of the cells increased in a dose-dependent manner, with the highest rate of about 67%. We evaluated the changes in the mRNA levels of genes related to the EA-mediated inhibition of cancer cell growth and initially verified the PI3K/PTEN/AKT/mTOR pathway as the pathway by which EA inhibits HeLa cell growth. We hope to provide a theoretical basis for the deep exploration and utilization of this functional food.

Proteins from Blackberry Seeds: Extraction, Osborne Isolate, Characteristics, Functional Properties, and Bioactivities.

Blackberry fruit contains high levels of nutrients and phenolic compounds. Blackberry pomace accounts for 20~30% of its whole fruit during processing and is generally treated as fertilizer. Blackberry pomace has many seeds that contain carbohydrates, polyphenols, flavonoids, pectin, protein, and other bioactive nutrients. However, its functional properties and seed protein compositions have not been reported. We used a single-factor experiment, response surface, and Osborne isolate method to extract protein isolate, albumin, globulin, glutelin, and prolamin from blackberry seeds for the first time and evaluated their characteristics and functional properties. Glutelin and protein isolate showed good water-holding capacity, emulsification, and foaming capacity, while albumin and globulin showed good oil-holding capacity and thermal stability. They were found to have good antioxidant activities that might be good DPPH free radical scavengers, especially prolamin, which has the lowest IC50 value (15.76 µg/mL). Moreover, globulin had the lowest IC50 value of 5.03 µg/mL against Hela cells, 31.82 µg/mL against HepG2 cells, and 77.81 µg/mL against MCF-7 cells and a high selectivity index (SI), which suggested globulin had better anti-cervical, antihepatoma, and anti-breast activity but relatively low cytotoxicity. These seed proteins may have great prospects for the development and application of food and drugs in the future.

Transcriptomic and Metabolomic Profiling Reveals the Variations in Carbohydrate Metabolism between Two Blueberry Cultivars.

Blueberry is a high-quality fruit tree with significant nutritional and economic value, but the intricate mechanism of sugar accumulation in its fruit remains unclear. In this study, the ripe fruits of blueberry cultivars 'Anna' and 'Misty' were utilized as experimental materials, and physiological and multi-omics methodologies were applied to analyze the regulatory mechanisms of the difference in sugar content between them. The results demonstrated that the 'Anna' fruit was smaller and had less hardness than the 'Misty' fruit, as well as higher sugar content, antioxidant capability, and lower active substance content. A total of 7067 differentially expressed genes (DEGs) (3674 up-regulated and 3393 down-regulated) and 140 differentially abundant metabolites (DAMs) (82 up-regulated and 58 down-regulated) were identified between the fruits of the two cultivars. According to KEGG analysis, DEGs were primarily abundant in phenylpropanoid synthesis and hormone signal transduction pathways, whereas DAMs were primarily enriched in ascorbate and aldarate metabolism, phenylpropanoid biosynthesis, and the pentose phosphate pathway. A combined multi-omics study showed that 116 DEGs and 3 DAMs in starch and sucrose metabolism (48 DEGs and 1 DAM), glycolysis and gluconeogenesis (54 DEGs and 1 DAM), and the pentose phosphate pathway (14 DEGs and 1 DAM) were significantly enriched. These findings suggest that blueberries predominantly increase sugar accumulation by activating carbon metabolism network pathways. Moreover, we identified critical transcription factors linked to the sugar response. This study presents new understandings regarding the molecular mechanisms underlying blueberry sugar accumulation and will be helpful in improving blueberry fruit quality through breeding.

The Extraction and High Antiproliferative Effect of Anthocyanin from Gardenblue Blueberry.

Blueberries are rich in flavonoids, anthocyanins, phenolic acids, and other bioactive substances. Anthocyanins are important functional components in blueberries. We collected 65 varieties of blueberries to investigate their nutritional and functional values. Among them, Gardenblue had the highest anthocyanin content, with 2.59 mg/g in fresh fruit. After ultrasound-assisted solvent extraction and macroporous resin absorption, the content was increased to 459.81 mg/g in the dried powder. Biological experiments showed that Gardenblue anthocyanins (L1) had antiproliferative effect on cervical cancer cells (Hela, 51.98 µg/mL), liver cancer cells (HepG2, 23.57 µg/mL), breast cancer cells (MCF-7, 113.39 µg/mL), and lung cancer cells (A549, 76.10 µg/mL), and no apparent toxic effects were indicated by methyl thiazolyl tetrazolium (MTT) assay, especially against HepG2 cells both in vitro and in vivo. After combining it with DDP (cisplatin) and DOX (doxorubicin), the antiproliferative effects were enhanced, especially when combined with DOX against HepG2 cells; the IC50 value was 0.02 µg/mL. This was further evidence that L1 could inhibit cell proliferation by inducing apoptosis. The detailed mechanism might be L1 interacting with DNA in an intercalation mode that changes or destroys DNA, causing apoptosis and inhibiting cell proliferation. The findings of this study suggest that L1 extract can be used as a functional agent against hepatoma carcinoma cells.

Small signaling peptides mediate plant adaptions to abiotic environmental stress.

MAIN CONCLUSION: Peptide-receptor complexes activate distinct downstream regulatory networks to mediate plant adaptions to abiotic environmental stress. Plants are constantly exposed to various adverse environmental factors; thus they must adjust their growth accordingly. Plants recruit small secretory peptides to adapt to these detrimental environments. These small peptides, which are perceived by their corresponding receptors and/or co-receptors, act as local- or long-distance mobile signaling molecules to establish cell-to-cell regulatory networks, resulting in optimal cellular and physiological outputs. In this review, we highlight recent advances on the regulatory role of small peptides in plant abiotic responses and nutrients signaling.

Absorbable suture anchor and knotless anchor techniques produced similar outcomes in arthroscopic anterior talofibular ligament repair.

PURPOSE: The purpose of this study is to compare absorbable suture anchor with knotless anchor techniques for arthroscopic anterior talofibular ligament (ATFL) repair. METHOD: A multicenter retrospective study was performed with 185 patients, who had undergone an arthroscopic ATFL repair procedure using absorbable suture anchor or knotless anchor between May 2017 and October 2019. The follow-up time was a minimum of 18 months. Karlsson-Peterson score, visual analogue scale (VAS), and Cumberland ankle instability tool (CAIT) were evaluated. The complications were also recorded. RESULTS: One hundred and seven patients underwent one absorbable suture anchor repair procedure (Group A [A]), and the other seventy-eight patients underwent one knotless anchor repair procedure (Group B [B]). At the final follow-up, both Karlsson-Peterson score (A, pre 61.0 ± 8.0 vs post 93.5 ± 5.3, P < 0.001; B, pre 59.5 ± 8.2 vs post 92.4 ± 6.3, P < 0.001), VAS score (A, pre 5.0 ± 1.3 vs post 0.5 ± 0.7, P < 0.001; B, pre 5.5 ± 1.2 vs post 0.9 ± 1.0, P < 0.001), and CAIT score (A, pre 53.1 ± 12.0 vs post 93.1 ± 6.6, P < 0.001; B, pre 51.6 ± 12.0 vs post 93.1 ± 6.5, P < 0.001) improved significantly in both groups. There was no significant difference between the two groups regarding the Karlsson-Peterson score (A, pre 61.0 ± 8.0 vs B, pre 59.5 ± 8.2, n.s; A, post 93.5 ± 5.3 vs B, post 92.4 ± 6.3, n.s), CAIT score (A, pre 53.1 ± 12.0 vs B, pre 51.6 ± 12.0, n.s; A, post 93.1 ± 6.6 vs B, post 93.1 ± 6.5, n.s) and the change ranges of VAS (A, 4.5 ± 1.0 vs B, 4.6 ± 1.2, n.s). Anchor complications were easier to occur in Group B (0/107 vs 6/78, P = 0.007). Knot irritation slightly increased in Group A (10/107 vs 0/78, P = 0.006). No significant difference was found regarding total complication rates (A, 10/107 vs B, 6/78, n.s). CONCLUSION: Absorbable suture anchor and knotless anchor for arthroscopic ATFL repair produced similar clinical outcomes. The ankle stability scores increased significantly in both groups. However, the knotless anchor has a higher risk to loosen, deviated direction or break, while the absorbable suture anchor still has a slim chance of knot irritation. LEVEL OF EVIDENCE: III.

Overexpression of RuFLS2 Enhances Flavonol-Related Substance Contents and Gene Expression Levels.

As an emerging third-generation fruit, blackberry has high nutritional value and is rich in polyphenols, flavonoids and anthocyanins. Flavonoid biosynthesis and metabolism is a popular research topic, but no related details have been reported for blackberry. Based on previous transcriptome data from this research group, two blackberry flavonol synthase genes were identified in this study, and the encoded proteins were subjected to bioinformatics analysis. RuFLS1 and RuFLS2 are both hydrophobic acidic proteins belonging to the 2OG-Fe(II) dioxygenase superfamily. RuFLS2 was expressed at 27.93-fold higher levels than RuFLS1 in red-purple fruit by RNA-seq analysis. Therefore, RuFLS2-overexpressing tobacco was selected for functional exploration. The identification of metabolites from transgenic tobacco showed significantly increased contents of flavonoids, such as apigenin 7-glucoside, kaempferol 3-O-rutinoside, astragalin, and quercitrin. The high expression of RuFLS2 also upregulated the expression levels of NtF3H and NtFLS in transgenic tobacco. The results indicate that RuFLS2 is an important functional gene regulating flavonoid biosynthesis and provides an important reference for revealing the molecular mechanism of flavonoid accumulation in blackberry fruit.

Phenolic profiles, antioxidant activity and inhibition of digestive enzymes of water caltrop pericarps.

BACKGROUND: Water caltrop (Trapa natans L.) is widely cultivated as a popular vegetable or fruit in Asian countries. In China, water caltrop pericarp is also used as a functional food to treat metabolic syndrome. However, the profiling of bioactive substances and their pharmacological activities in different water caltrop varieties remains to be investigated. In the present study, three varieties of water caltrop pericarps collected from 13 origins in China were analyzed for their phenolic substances. To investigate the pharmacological activities, samples were tested for their free radical scavenging capacity and inhibitory potency against α-glucosidase and pancreatic lipase. RESULTS: In total, 46 phenolic compounds were identified in the ethanol extract of water caltrop pericarp using a liquid chromatography-quadrupole time of flight-tandem mass spectrometry method, most of which were hydrolyzable tannins. Two cultivated varieties samples exhibited a relatively higher phenolic content and stronger antioxidant and inhibitory activities against α-glucosidase and pancreatic lipase compared to those from the wild variety. Correlation analysis between phenolic contents and biological activities suggested that phenolic compounds exhibited potential free radical scavenging capacity, α-glucosidase and pancreatic lipase inhibitory activities. CONCLUSION: It is concluded that the phenolic compounds of water caltrop pericarp are promising sources of natural antioxidants, α-glucosidase and pancreatic lipase inhibitors. © 2021 Society of Chemical Industry.

Arthroscopic Suture-Bridge Repair Technique for an Avulsion of the Talar Insertion of the Anterior Talofibular Ligament.

There are several studies about arthroscopic repair techniques for the lesion of the anterior talofibular ligament. However, the research concentrating on the avulsion of the talar insertion of the anterior talofibular ligament is very rare. Among 122 patients who suffered from recurrent ankle sprain and underwent arthroscopic anterior talofibular ligament repair from October 2016 to January 2019 in our hospital, 11 patients with an avulsion of the talar insertion of this ligament were diagnosed and then treated with the arthroscopic suture-bridge repair technique in the present study. The clinical outcomes were assessed using the Karlsson-Peterson score, Ankle and hindfoot score by American Orthopedic of Foot and Ankle Society, Sefton articular stability scale and Visual Analogue Scale. The complications were recorded at the time of observation. The median value of the follow-up time was 30 (range 18-36) months. At the final follow-up, the median value of the Karlsson-Peterson score, American Orthopedic of Foot and Ankle Society ankle-hindfoot score, and Visual Analogue Scale score was 90, 90, 1, respectively. Based on the Sefton stability scale, 10 cases were in the excellent or good category. No wound infections and no neurovascular injuries were encountered, also no case required revision surgery. Only 1 patient complained about mild local irritation at the knotless anchor site. The arthroscopic suture-bridge technique could be suitable for treatment of an avulsion of the talar insertion of the anterior talofibular ligament due to satisfactory activity recovery and few complications.

In Colorectal Cancer Cells With Mutant KRAS, SLC25A22-Mediated Glutaminolysis Reduces DNA Demethylation to Increase WNT Signaling, Stemness, and Drug Resistance.

BACKGROUND & AIMS: Mutant KRAS promotes glutaminolysis, a process that uses steps from the tricarboxylic cycle to convert glutamine to α-ketoglutarate and other molecules via glutaminase and SLC25A22. This results in inhibition of demethylases and epigenetic alterations in cells that increase proliferation and stem cell features. We investigated whether mutant KRAS-mediated glutaminolysis affects the epigenomes and activities of colorectal cancer (CRC) cells. METHODS: We created ApcminKrasG12D mice with intestine-specific knockout of SLC25A22 (ApcminKrasG12DSLC25A22fl/fl mice). Intestine tissues were collected and analyzed by histology, immunohistochemistry, and DNA methylation assays; organoids were derived and studied for stem cell features, along with organoids derived from 2 human colorectal tumor specimens. Colon epithelial cells (1CT) and CRC cells (DLD1, DKS8, HKE3, and HCT116) that expressed mutant KRAS, with or without knockdown of SLC25A22 or other proteins, were deprived of glutamine or glucose and assayed for proliferation, colony formation, glucose or glutamine consumption, and apoptosis; gene expression patterns were analyzed by RNA sequencing, proteins by immunoblots, and metabolites by liquid chromatography-mass spectrometry, with [U-13C5]-glutamine as a tracer. Cells and organoids with knocked down, knocked out, or overexpressed proteins were analyzed for DNA methylation at CpG sites using arrays. We performed immunohistochemical analyses of colorectal tumor samples from 130 patients in Hong Kong (57 with KRAS mutations) and Kaplan-Meier analyses of survival. We analyzed gene expression levels of colorectal tumor samples in The Cancer Genome Atlas. RESULTS: CRC cells that express activated KRAS required glutamine for survival, and rapidly incorporated it into the tricarboxylic cycle (glutaminolysis); this process required SLC25A22. Cells incubated with succinate and non-essential amino acids could proliferate under glutamine-free conditions. Mutant KRAS cells maintained a low ratio of α-ketoglutarate to succinate, resulting in reduced 5-hydroxymethylcytosine-a marker of DNA demethylation, and hypermethylation at CpG sites. Many of the hypermethylated genes were in the WNT signaling pathway and at the protocadherin gene cluster on chromosome 5q31. CRC cells without mutant KRAS, or with mutant KRAS and knockout of SLC25A22, expressed protocadherin genes (PCDHAC2, PCDHB7, PCDHB15, PCDHGA1, and PCDHGA6)-DNA was not methylated at these loci. Expression of the protocadherin genes reduced WNT signaling to ß-catenin and expression of the stem cell marker LGR5. ApcminKrasG12DSLC25A22fl/fl mice developed fewer colon tumors than ApcminKrasG12D mice (P < .01). Organoids from ApcminKrasG12DSLC25A22fl/fl mice had reduced expression of LGR5 and other markers of stemness compared with organoids derived from ApcminKrasG12D mice. Knockdown of SLC25A22 in human colorectal tumor organoids reduced clonogenicity. Knockdown of lysine demethylases, or succinate supplementation, restored expression of LGR5 to SLC25A22-knockout CRC cells. Knockout of SLC25A22 in CRC cells that express mutant KRAS increased their sensitivity to 5-fluorouacil. Level of SLC25A22 correlated with levels of LGR5, nuclear ß-catenin, and a stem cell-associated gene expression pattern in human colorectal tumors with mutations in KRAS and reduced survival times of patients. CONCLUSIONS: In CRC cells that express activated KRAS, SLC25A22 promotes accumulation of succinate, resulting in increased DNA methylation, activation of WNT signaling to ß-catenin, increased expression of LGR5, proliferation, stem cell features, and resistance to 5-fluorouacil. Strategies to disrupt this pathway might be developed for treatment of CRC.

FK506 ameliorates osteoporosis caused by osteoblast apoptosis via suppressing the activated CaN/NFAT pathway during oxidative stress.

OBJECTIVE: Osteoporosis is affecting the health of postmenopausal women in the world. In case of that, we explored whether FK-506 could ameliorate osteoporosis by inhibiting the activated CaN/NFAT pathway during oxidative stress. METHODS: First, the castrated rat model is constructed through the bilateral ovariectomy. Hologic Discovery (S/N 80347) dual-energy X-ray absorptiometry assessed bone mineral density (BMD) implemented at left femur of rats. Next, hematoxylin-eosin (H&E) staining observed and calculated the changes of bone trabecular, mean trabecular plate separation (Tb.Sp), mean trabecular plate thickness (Tb.Th), and bone volume fraction (BV/TV). Then, CCK-8 assay, TUNEL assay, ALP kit and alizarin red staining detected the viability, apoptosis, alkaline phosphatase (ALP) activity, and capacity of mineralization respectively. At last, commercially available kits detected the levels of ROS and SOD in transfected MC3T3-E1 cells and bone tissues, and Western blot analysis detected proteins related to apoptosis and CaN/NFAT pathway. RESULTS: FK-506 increased the BMD and changes of bone trabecular in female castrated rats. FK-506 inhibited the oxidative stress and apoptosis by suppressing the activated CaN/NFAT pathway. Low dose of FK-506 improved the viability, ALP activity, and mineralization capacity. What's more, it suppressed the apoptosis of H2O2-induced MC3T3-E1 cells, which was deteriorated by the high dose of FK-506. Briefly, low dose of FK-506 inhibited the oxidative stress by suppressing the activated CaN/NFAT pathway, while high dose of that further inhibited the oxidative stress by suppressing the CaN/NFAT pathway. CONCLUSION: FK-506 ameliorates osteoporosis resulted from osteoblastic apoptosis which caused by suppressing the activated CaN/NFAT pathway during oxidative stress.

Autopsy and statistical evidence of disturbed hemostasis progress in COVID-19: medical records from 407 patients.

BACKGROUND: The progression of coagulation in COVID-19 patients with confirmed discharge status and the combination of autopsy with complete hemostasis parameters have not been well studied. OBJECTIVE: To clarify the thrombotic phenomena and hemostasis state in COVID-19 patients based on epidemiological statistics combining autopsy and statistical analysis. METHODS: Using autopsy results from 9 patients with COVID-19 pneumonia and the medical records of 407 patients, including 39 deceased patients whose discharge status was certain, time-sequential changes in 11 relevant indices within mild, severe and critical infection throughout hospitalization according to the Chinese National Health Commission (NHC) guidelines were evaluated. Statistical tools were applied to calculate the importance of 11 indices and the correlation between those indices and the severity of COVID-19. RESULTS: At the beginning of hospitalization, platelet (PLT) counts were significantly reduced in critically ill patients compared with severely or mildly ill patients. Blood glucose (GLU), prothrombin time (PT), activated partial thromboplastin time (APTT), and D-dimer levels in critical patients were increased compared with mild and severe patients during the entire admission period. The International Society on Thrombosis and Haemostasis (ISTH) disseminated intravascular coagulation (DIC) score was also high in critical patients. In the relatively late stage of nonsurvivors, the temporal changes in PLT count, PT, and D-dimer levels were significantly different from those in survivors. A random forest model indicated that the most important feature was PT followed by D-dimer, indicating their positive associations with disease severity. Autopsy of deceased patients fulfilling diagnostic criteria for DIC revealed microthromboses in multiple organs. CONCLUSIONS: Combining autopsy data, time-sequential changes and statistical methods to explore hemostasis-relevant indices among the different severities of the disease helps guide therapy and detect prognosis in COVID-19 infection.

Regulation of Apoptosis and Inflammatory Response in Interleukin-1ß-Induced Nucleus Pulposus Cells by miR-125b-5p Via Targeting TRIAP1.

The aim of the present study was to determine the function of microRNA (miR)-125b-5p in lumbar disc degeneration (LDD). Nucleus pulposus (NP) cells were stimulated with 10 ng/ml IL-1ß for 24 h to establish an LDD model. Reverse transcription-quantitative PCR was used to assess miR-125b-5p levels in human lumbar degenerative NP samples and IL-1ß-treated NP cells. An interaction between miR-125b-5p and TP53-regulated inhibitor of apoptosis 1 (TRIAP1) was revealed by TargetScan 7.1 and dual-luciferase reporter assay. Protein levels of pro-inflammatory factors were determined using ELISA. Cell viability and apoptosis were evaluated by MTT and flow cytometry analysis, respectively. miR-125b-5p was markedly upregulated in both human lumbar degenerative NP specimens and IL-1ß-treated NP cells. TRIAP1, which directly targets miR-125b-5p, was markedly downregulated in human lumbar degenerative NP specimens and IL-1ß-treated NP cells. The levels of TNF-α and IL-6 were inhibited in IL-1ß-treated NP cells transfected with miR-125b-5p inhibitor. Moreover, miR-125b-5p inhibitor increased NP cell viability, prevented apoptosis and repressed the apoptotic peptidase activating factor 1/caspase 9 pathway in IL-1ß-treated NP cells. Thus, the present findings suggested that miR-125b-5p could regulate LDD by adjusting NP cell apoptosis and inflammatory responses via TRIAP1.

Selection and Validation of Candidate Reference Genes for Gene Expression Analysis by RT-qPCR in Rubus.

Due to the lack of effective and stable reference genes, studies on functional genes in Rubus, a genus of economically important small berry crops, have been greatly limited. To select the best internal reference genes of different types, we selected four representative cultivars of blackberry and raspberry (red raspberry, yellow raspberry, and black raspberry) as the research material and used RT-qPCR technology combined with three internal stability analysis software programs (geNorm, NormFinder, and BestKeeper) to analyze 12 candidate reference genes for the stability of their expression. The number of most suitable internal reference genes for different cultivars, tissues, and fruit developmental stages of Rubus was calculated by geNorm software to be two. Based on the results obtained with the three software programs, the most stable genes in the different cultivars were RuEEF1A and Ru18S. Finally, to validate the reliability of selected reference genes, the expression pattern of the RuCYP73A gene was analyzed, and the results highlighted the importance of appropriate reference gene selection. RuEEF1A and Ru18S were screened as reference genes for their relatively stable expression, providing a reference for the further study of key functional genes in blackberry and raspberry and an effective tool for the analysis of differential gene expression.

Epigenomic biomarkers for prognostication and diagnosis of gastrointestinal cancers.

Altered epigenetic regulation is central to many human diseases, including cancer. Over the past two decade, major advances have been made in our understanding of the role of epigenetic alterations in carcinogenesis, particularly for DNA methylation, histone modifications and non-coding RNAs. Aberrant hypermethylation of DNA at CpG islands is a well-established phenomenon that mediates transcriptional silencing of tumor suppressor genes, and it is an early event integral to gastrointestinal cancer development. As such, detection of aberrant DNA methylation is being developed as biomarkers for prognostic and diagnostic purposes in gastrointestinal cancers. Diverse tissue types are suitable for the analyses of methylated DNA, such as tumor tissues, blood, plasma, and stool, and some of these markers are already utilized in the clinical setting. Recent advances in the genome-wide epigenomic approaches are enabling the comprehensive mapping of the cancer methylome, thus providing new avenues for mining novel biomarkers for disease prognosis and diagnosis. Here, we review the current knowledge on DNA methylation biomarkers for the prognostication and non-invasive diagnosis of gastrointestinal cancers and highlight their clinical application.

tiRNAs: A novel class of small noncoding RNAs that helps cells respond to stressors and plays roles in cancer progression.

tRNA-derived stress-induced RNAs (tiRNAs), important components of tRNA-derived fragments, are gaining popularity for their functions as small noncoding RNAs involved in cancer progression. Under cellular stress, tiRNAs are generated when mature tRNA is specifically cleaved by angiogenin and suggested to act as transducers or effectors involved in cellular stress responses. tiRNAs facilitate cells to respond to stresses mainly via reprogramming translation, inhibiting apoptosis, degrading mRNA, and generating stress granules. This review introduces the cellular biogenesis, molecular mechanisms, and biological roles of tiRNAs in stress response and disease regulation. A better understanding of their roles in regulating cancer may provide novel biomarkers or therapeutic targets for diagnosis and treatment.