Identification of diagnostic biomarkers of rheumatoid arthritis based on machine learning-assisted comprehensive bioinformatics and its correlation with immune cells.

Background: Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease characterized by inflammatory cell infiltration, which can lead to chronic disability, joint destruction and loss of function. At present, the pathogenesis of RA is still unclear. The purpose of this study is to explore the potential biomarkers and immune molecular mechanisms of rheumatoid arthritis through machine learning-assisted bioinformatics analysis, in order to provide reference for the early diagnosis and treatment of RA disease. Methods: RA gene chips were screened from the public gene GEO database, and batch correction of different groups of RA gene chips was performed using Strawberry Perl. DEGs were obtained using the limma package of R software, and functional enrichment analysis such as gene ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), disease ontology (DO), and gene set (GSEA) were performed. Three machine learning methods, least absolute shrinkage and selection operator regression (LASSO), support vector machine recursive feature elimination (SVM-RFE) and random forest tree (Random Forest), were used to identify potential biomarkers of RA. The validation group data set was used to verify and further confirm its expression and diagnostic value. In addition, CIBERSORT algorithm was used to evaluate the infiltration of immune cells in RA and control samples, and the correlation between confirmed RA diagnostic biomarkers and immune cells was analyzed. Results: Through feature screening, 79 key DEGs were obtained, mainly involving virus response, Parkinson's pathway, dermatitis and cell junction components. A total of 29 hub genes were screened by LASSO regression, 34 hub genes were screened by SVM-RFE, and 39 hub genes were screened by Random Forest. Combined with the three algorithms, a total of 12 hub genes were obtained. Through the expression and diagnostic value verification in the validation group data set, 7 genes that can be used as diagnostic biomarkers for RA were preliminarily confirmed. At the same time, the correlation analysis of immune cells found that γδT cells, CD4+ memory activated T cells, activated dendritic cells and other immune cells were positively correlated with multiple RA diagnostic biomarkers, CD4+ naive T cells, regulatory T cells and other immune cells were negatively correlated with multiple RA diagnostic biomarkers. Conclusions: The results of novel characteristic gene analysis of RA showed that KYNU, EVI2A, CD52, C1QB, BATF, AIM2 and NDC80 had good diagnostic and clinical value for the diagnosis of RA, and were closely related to immune cells. Therefore, these seven DEGs may become new diagnostic markers and immunotherapy markers for RA.

What role does the seed coat play during symbiotic seed germination in orchids: an experimental approach with Dendrobium officinale.

BACKGROUND: Orchids require specific mycorrhizal associations for seed germination. During symbiotic germination, the seed coat is the first point of fungal attachment, and whether the seed coat plays a role in the identification of compatible and incompatible fungi is unclear. Here, we compared the effects of compatible and incompatible fungi on seed germination, protocorm formation, seedling development, and colonization patterns in Dendrobium officinale; additionally, two experimental approaches, seeds pretreated with NaClO to change the permeability of the seed coat and fungi incubated with in vitro-produced protocorms, were used to assess the role of seed coat played during symbiotic seed germination. RESULTS: The two compatible fungi, Tulasnella sp. TPYD-2 and Serendipita indica PI could quickly promote D. officinale seed germination to the seedling stage. Sixty-two days after incubation, 67.8 ± 5.23% of seeds developed into seedlings with two leaves in the PI treatment, which was significantly higher than that in the TPYD-2 treatment (37.1 ± 3.55%), and massive pelotons formed inside the basal cells of the protocorm or seedlings in both compatible fungi treatments. In contrast, the incompatible fungus Tulasnella sp. FDd1 did not promote seed germination up to seedlings at 62 days after incubation, and only a few pelotons were occasionally observed inside the protocorms. NaClO seed pretreatment improved seed germination under all three fungal treatments but did not improve seed colonization or promote seedling formation by incompatible fungi. Without the seed coat barrier, the colonization of in vitro-produced protocorms by TPYD-2 and PI was slowed, postponing protocorm development and seedling formation compared to those in intact seeds incubated with the same fungi. Moreover, the incompatible fungus FDd1 was still unable to colonize in vitro-produced protocorms and promote seedling formation. CONCLUSIONS: Compatible fungi could quickly promote seed germination up to the seedling stage accompanied by hyphal colonization of seeds and formation of many pelotons inside cells, while incompatible fungi could not continuously colonize seeds and form enough protocorms to support D. officinale seedling development. The improvement of seed germination by seed pretreatment may result from improving the seed coat hydrophilicity and permeability, but seed pretreatment cannot change the compatibility of a fungus with an orchid. Without a seed coat, the incompatible fungus FDd1 still cannot colonize in vitro-produced protocorms or support seedling development. These results suggest that seed coats are not involved in symbiotic germination in D. officinale.

Compatible and Incompatible Mycorrhizal Fungi With Seeds of Dendrobium Species: The Colonization Process and Effects of Coculture on Germination and Seedling Development.

Orchids highly rely on mycorrhizal fungi for seed germination, and compatible fungi could effectively promote germination up to seedlings, while incompatible fungi may stimulate germination but do not support subsequent seedling development. In this study, we compared the fungal colonization process among two compatible and two incompatible fungi during seed germination of Dendrobium officinale. The two compatible fungi, i.e., Tulasnella SSCDO-5 and Sebacinales LQ, originally from different habitats, could persistently colonize seeds and form a large number of pelotons continuously in the basal cells, and both fungi promoted seed germination up to seedling with relative effectiveness. In contrast, the two incompatible fungi, i.e., Tulasnella FDd1 and Tulasnella AgP-1, could not persistently colonize seeds. No pelotons in the FDd1 treatment and only a few pelotons in the AgP-1 treatment were observed; moreover, no seedlings were developed at 120 days after incubation in either incompatible fungal treatment. The pattern of fungal hyphae colonizing seeds was well-matched with the morphological differentiation of seed germination and seedling development. In the fungal cocultural experiments, for both orchids of D. officinale and Dendrobium devonianum, cocultures had slightly negative effects on seed germination, protocorm formation, and seedling formation compared with the monocultures with compatible fungus. These results provide us with a better understanding of orchid mycorrhizal interactions; therefore, for orchid conservation based on symbiotic seed germination, it is recommended that a single, compatible, and ecological/habitat-specific fungus can be utilized for seed germination.

Oxidation pretreatment by calcium hypochlorite to improve the sensitivity of enzyme inhibition-based detection of organophosphorus pesticides.

BACKGROUND: Enzyme inhibition-based detection is the most widely used method for rapid detection of organophosphorus pesticides (OPs) in food and agricultural products. However, the accuracy of the method is negatively affected by low inhibitory activities of OPs with PS moiety on acetylcholinesterase. RESULTS: We demonstrated that oxidation pretreatments with bromine, hydrogen peroxide, or calcium hypochlorite significantly enhanced the enzyme inhibitory activities of these OPs. Especially, calcium hypochlorite (0.05%) pretreatment converted the PS moiety in OPs to PO and produced the most potent and steady inhibitory effect on the enzyme. This, in turn, resulted in a dramatic increase in the sensitivity of enzyme inhibition-based detection of these OPs by as much as 2 to 7 orders of magnitude. Importantly, this enhanced detection of OPs was validated in various vegetable samples. CONCLUSION: Our findings provide a solid basis to use calcium hypochlorite pretreatment for the improved detection of OPs by the enzyme inhibition-based method. © 2017 Society of Chemical Industry.

A systematic determination of polyphenols constituents and cytotoxic ability in fruit parts of pomegranates derived from five Chinese cultivars.

Plant polyphenols derived from pomegranates are natural health-promoting components, and their bioactivities are well proved. However, the systematic studies of polyphenols constituents and cytotoxic ability in fruit parts of pomegranates derived from different Chinese cultivars have not been studied yet. In this report, a validated and sensitive HPLC-DAD method and fluorescence spectrophotometric method was established for quantitative analysis of four polyphenols and total phenolic content (TPC) in fruit parts of pomegranates (including peels, flesh, seeds, juices and leaves) derived from five Chinese cultivars, respectively. HPLC analysis was performed on the YMC ODS-A C18 column with gradient elution of MeOH and 0.1 % TFA. Four polyphenols including gallic acid, ellagic acid, punicalagin A&B and punicalin A&B exhibited satisfactory linearity in the concentration ranges of 20-320, 39-624, 74-1184 and 38-608 µg/mL, respectively. The results demonstrated that the amounts of TPC and four polyphenols in different fruit parts of pomegranates varied significantly. Peels of Sour-YRP possessed the highest content of punicalagin A&B (125.23 mg/g), whereas other three polyphenols exhibited only trace. Among the five Chinese cultivars, Sour-YRP contained the highest content of TPC (688.61 mg/g) and could be considered as the desirable botanical source to obtain polyphenols. It is also discovered that low-maturity pomegranate might possessed much higher TPC than high-maturity pomegranate. The optimized HPLC-DAD method could be used for quality control of different pomegranates by identification and quantification of its main polyphenolic components. Furthermore, the in vitro cytotoxicity of different pomegranates fruit parts to cancer cells was evaluated. We discovered that peels and flesh extract of Sour-YRP significantly inhibited the proliferation of HepG2 and Hela cancer cells lines. The results of this work are promising for further investigation and development of pomegranates as therapeutic agent for the treatment of cancer.

An integrated molecular docking and rescoring method for predicting the sensitivity spectrum of various serine hydrolases to organophosphorus pesticides.

BACKGROUND: The enzymatic chemistry method is currently the most widely used method for the rapid detection of organophosphorus (OP) pesticides, but the enzymes used, such as cholinesterases, lack sufficient sensitivity to detect low concentrations of OP pesticides present in given samples. Serine hydrolase is considered an ideal enzyme source in seeking high-sensitivity enzymes used for OP pesticide detection. However, it is difficult to systematically evaluate sensitivities of various serine hydrolases to OP pesticides by in vitro experiments. This study aimed to establish an in silico method to predict the sensitivity spectrum of various serine hydrolases to OP pesticides. RESULTS: A serine hydrolase database containing 219 representative serine hydrolases was constructed. Based on this database, an integrated molecular docking and rescoring method was established, in which the AutoDock Vina program was used to produce the binding poses of OP pesticides to various serine hydrolases and the ID-Score method developed recently by us was adopted as a rescoring method to predict their binding affinities. In retrospective case studies, this method showed good performance in predicting the sensitivities of known serine hydrolases to two OP pesticides: paraoxon and diisopropyl fluorophosphate. The sensitivity spectrum of the 219 collected serine hydrolases to 37 commonly used OP pesticides was finally obtained using this method. CONCLUSION: Overall, this study presented a promising in silico tool to predict the sensitivity spectrum of various serine hydrolases to OP pesticides, which will help in finding high-sensitivity serine hydrolases for OP pesticide detection.

[Preliminary processing, processing and usage of Dendrobii Caulis in history].

On account of the dense cuticles of the fresh stem and the light, hard and pliable texture of the dried stem, Dendrobii Caulis is difficult to dry or pulverize. So, it is very important to the ancient doctors that Dendrobii Caulis should be properly treated and applied to keep or evoke its medicinal effects. The current textual research results about the preliminary processing, processing and usage methods of Dendrobii Caulis showed that: (1) In history the clinical use of fresh or processed Dendrobii Caulis as teas and tinctures were very common. (2) Its roots and rhizomes would be removed before using. (3) Some ancillary approaches were applied to shorten drying times, such as rinsing with boiling mulberry-ash soup, washing or soaking with liquor, mixing with rice pulp and then basking, etc. (4) According to the ancients knowledge, the sufficient pulverization, by means of slicing, rasping, hitting or pestling techniques, was necessary for Dendrobii Caulis to take its effects. (5) The heat processing methods for Dendrobii Caulis included stir-baking, stir-frying, steaming, decocting and stewing techniques, usually with liquor as an auxiliary material. Among above mentioned, steaming by pretreating with liquor was most commonly used, and this scheme was colorfully drawn in Bu Yi Lei Gong Pao Zhi Bian Lan (Ming Dynasty, 1591 CE) ; moreover, decocting in advance or long-time simmering so as to prepare paste products were recommended in the Qing Dynasty. (6) Some different processing programs involving stir-baking with grit, air-tightly baking with ondol (Kangs), fumigating with sulfur, which appeared in modern times and brought attractive outward appearance of the drug, went against ancients original intentions of ensuring drug efficacy.

Actinopolyspora lacussalsi sp. nov., an extremely halophilic actinomycete isolated from a salt lake.

A novel halophilic, filamentous actinobacterium, designated strain TRM 40139(T), was isolated from a hypersaline habitat in Xinjiang Province, north-west China. Its taxonomic status was determined using a polyphasic approach. Phylogenetic analysis based on the almost-complete 16S rRNA gene sequence of the strain showed that it formed a well-separated sub-branch within the radiation of the genus Actinopolyspora and the organism was related most closely to the type strains of Actinopolyspora alba (97.6 % similarity), Actinopolyspora xinjiangensis (97.6 %) and Actinopolyspora erythraea (97.1 %). However, it had relatively lower mean DNA-DNA relatedness values with the above strains (36.4, 31.3 and 26.1 %, respectively). Optimal growth occurred at 35 °C, at pH 7.0 and in the presence of 12 % (w/v) NaCl. The whole-cell sugar pattern consisted of xylose, glucose, ribose and arabinose. The major fatty acids were iso-C16 : 0 (28.0 %) and anteiso-C17 : 0 (27.6 %). The diagnostic phospholipids detected were diphosphatidylglycerol, phosphatidylglycerol, phosphatidylcholine, phosphatidylinositol and two unknown phospholipids. The predominant menaquinones were MK-9(H4) (49.8 %) and MK-10(H4) (24.2 %). The G+C content of the genomic DNA was 66.4 mol%. Strain TRM 40139(T) therefore represents a novel species of the genus Actinopolyspora, for which the name Actinopolyspora lacussalsi sp. nov. is proposed. The type strain is TRM 40139(T) (= KCTC 19657(T) = CCTCC AA 2012020(T)).

Myceligenerans salitolerans sp. nov., a halotolerant actinomycete isolated from a salt lake in Xinjiang, China.

A halotolerant actinomycete strain, designated XHU 5031(T), was isolated from a salt lake in Xinjiang Province, northwest China. Its taxonomic status was determined using a polyphasic approach. Phylogenetic analysis based on 16S rRNA gene sequences revealed that the organism was most closely related to Myceligenerans xiligouense DSM 15700(T) (98.4 %), Myceligenerans halotolerans XJEEM 11063(T) (98.0 %) and Myceligenerans crystallogenes DSM 17134(T) (97.5 %). However, it had relatively low values for DNA-DNA relatedness with the above strains (46.2, 39.4 and 36.5 %, respectively). The peptidoglycan type was A4α. This organism contained glucose, mannose and galactose as the major whole cell sugars. The predominant menaquinone was MK-9(H(4)). The major fatty acids were iso-C(15:0,) anteiso-C(15:0) and iso-C(16:0). The polar lipids detected were diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylinositol (PI), one unknown phospholipid (PL) and two unknown glycolipids (GL). The G+C content of genomic DNA was 71.2 mol %. Phenotypic data clearly distinguished the isolate from its closest relatives. The combined genotypic and phenotypic data indicated that the isolate XHU 5031(T) represented a novel species of the genus Myceligenerans. The proposed name for this organism is Myceligenerans salitolerans sp. nov., with type strain XHU 5031(T) (=KCTC 29128(T) = CCTCC AB 2012908(T)).

Assessment of iron bioavailability in ten kinds of Chinese wheat flours using an in vitro digestion/Caco-2 cell model.

OBJECTIVE: To compare iron bioavailability (Fe BV) from ten selected kinds of Chinese wheat flours in order to provide scientific basis for further human trials and enable plant breeding programs to screen biofortified wheat cultivars. METHODS: An in vitro digestion/Caco-2 cell model was used to assess Fe BV of ten flour samples from six leading Chinese wheat cultivars and the stability of Fe BV in one cultivar was studied across three growing environments. RESULTS: Significant differences were observed in both Fe BV and Fe bioavailability per gram of food (Fe BVPG) among cultivars (P<0.01) grown at the same location with the same flour extraction rate. Zhongyou 9507 and Jingdong 8 had Fe BV 37%-54% and Fe BVPG 103%-154% higher than the reference control. In the Anyang environment, Zhongyou 9507 had a higher wheat flour-Fe level and Fe BVPG. Differences in Fe BV were detected in cultivars with different flour extraction rates. CONCLUSION: Zhongyou 9507 and Jingdong 8 were identified as the most promising cultivars for further evaluation of efficacy by using human subjects. The growing environments had no effect on Fe BV, but did have a significant effect on Fe BVPG. Fe bioavailabilities in low-extraction (40%) flours were higher than those in high-extraction (78%) flours.

[Analysis of correlation of local GC level in human protein coding genes].

GC level is an important feature of genomic composition, which significantly improve our understanding of structure, function and evolution of genes. In this paper, the nonredundant DNA sequence of 7,992 human protein coding genes were retrieved from public database and the local GC level of different sequence regions and correlation between GC levels were analyzed.. The results showed that the GC levels of different sequence regions were strikingly nonuniform. 5' untranslated regions were of richest GC, with average GC content being 62.5%. 3'-untranslated regions were of poorest GC, with average GC content being 43.97%. GC contents of 3' flanking sequences profoundly matched the GC levels of DNA large fragments where the genes were located. Although the GC contents of open reading frames (ORFs) were higher than that of intron, 3' non-translated region and 3' flanking sequences, high correlation existed among the GC contents of the four regions. Average GC content of the third codon position (GC3) was 58.9%, higher than that of the fist and second position, and showed high correlation to GC contents of ORFs, with correlation coefficients being 0.91, besides of its significant association with GC contents of intron, 3'-untranslated region and 3' flanking sequences. Moreover, the linear regression of GC3 against GC contents of 3' flanking sequences yielded a slope of 1.25. Thus, GC3 was a sensitive indicator for GC change of local genome. As for 5' flanking sequences, 5' untranslated regions, fist and second codon position, however, their GC level exhibited weaker correlation with that of other regions. These results suggest that the third codon positions, introns, 3'-untranslated regions and 3' flanking sequences may evolve similarly while first and second codon positions, 5' flanking sequences and 5' untranslated region were expected to bear more selective stress for holding their functions.

[Effects of exogenerous nucleotides on the apoptosis of intestinal epithelial cells IEC-6].

OBJECTIVE: To investigate the effects of exogenous nucleotides on apoptosis of a normal rat small intestinal epithelial cell line, IEC-6. METHODS: Cultured IEC-6 cells were treated by four kinds of monophosphate nucleotides and their mixture prepared according to their composition in human milk, then the cell apoptosis was determined by flow cytometry measurement, morphologic characterization, and electron-microscope observation. RESULTS: IEC-6 cells treated with AMP or GMP showed a apotosis peak in flow cytometry measurement, but only AMP produce typical apoptosis characteristics in electron-microscope observation. Pyrimidine nucleotides (UMP and CMP)and nucleotides mixture could not induce apoptosis. However, UMP could significantly eliminate the apoptosis-inducing effects of AMP or GMP. CONCLUSION: Purine nucleotides induce apoptosis of IEC-6, inducing effects of purine nucleotides. pyrimidine nucleotides UMP could abolish the apoptosis-inducing effects of purine nucleotides.

[Experimental studies on male reproductive toxicity of bisphenol A in vitro and vivo].

OBJECTIVE: To explore the effects of Bisphenol A in adult rats and its possible mechanisms. METHODS: BPA (in corn oil) was administered orally to 9-week-old male Sprague-Dawley rats for 14 days (0, 1 and 5 g/kg bw), and incubated primary Sertoli cells from pubertal SD rats with 0, 10(-7), 10(-6), 10(-5), 10(-4) mol/L BPA. RESULTS: After oral administration, a significant decrease in right testis weight was observed in 5 g/kg dose group, but not in the 1 g/kg bw dose group. Germ cells were detached from basement membrane of seminiferous tubules and Sertoli cells in BPA-treated groups. Administration of BPA at 1 g/kg bw and 5 g/kg bw produced both nucleus pycnosis and vacuolized nucleus in germ cells and Sertoli cells. A marked loss in vimentin staining in Sertoli cells from testis of BPA-treated rats was detected. No change in levels of serum estradiol and testosterone was observed after two-week exposure to BPA. In Sertoli cell primary culture, BPA destroyed the cytoskeleton and cell-cell junctions, and elongated Sertoli cells. CONCLUSION: These results suggest that BPA may injure reproductive function of male rats by destroying the cytoskeleton and changing the form of Sertoli cells.