[Transcriptome data mining combined with clinical and animal experiments for identification of syndrome element marker genes during entire course of steroid-induced osteonecrosis of femoral head].

A research strategy combining transcriptome data mining and experimental verification was adopted to identify the marker genes characterizing the syndrome elements of phlegm, stasis, and deficiency in steroid-induced osteonecrosis of the femoral head(SONFH). Firstly, the common differentially expressed gene sets of SONFH with the syndromes of phlegm-stasis obstructing collaterals, vessel obstruction, and liver-kidney deficiency were obtained from the clinical transcriptomic analysis of a previous study. The differential expression trend analysis and functional gene mining were then employed to predict the candidate marker gene sets representing phlegm, stasis, and deficiency. The whole blood samples from SONFH patients, whole blood samples from SONFH rats, and affected femoral head tissue samples were collected for qPCR, which aimed to determine the expression levels of the candidate marker genes mentioned above. Furthermore, the receiver operating characteristic curve(ROC) was established to objectively evaluate the syndrome differentiation effectiveness of the candidate marker genes mentioned above. The transcriptome data analysis results showed that the candidate marker genes for phlegm was ELOVL fatty acid elongase 6(ELOVL6), and those for stasis were ankyrin 1(ANK1), glycophorin A/B(GYPA/B), and Rh-associated glycoprotein(RHAG). The candidate marker genes for deficiency were solute carrier family 2 member 1(SLC2A1) and stomatin(STOM). The qPCR results showed that compared with that in the non-SONFH group, ELOVL6 had the lowest expression level in the peripheral blood of the SONFH patients with the syndrome of phlegm-stasis obstructing collaterals(P<0.05). Compared with that in the normal control group, ELOVL6 had the lowest expression level in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 4 weeks(P<0.01), and it showed better syndrome differentiation effectiveness of rats modeled for 4 weeks(AUC=0.850, P=0.006) than at other modeling time points(8, 12, 16, and 21 weeks, AUC of 0.689, 0.766, 0.588, and 0.662, respectively). Compared with that in the non-SONFH group, the expression levels of ANK1, GYPA, and RHAG were the lowest in the peripheral blood of SONFH patients with the vessel obstruction syndrome(P<0.05). The expression levels of the three genes were the lowest in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 12 weeks(P<0.05, P<0.01), and their syndrome differentiation effectiveness in the rats modeled for 12 weeks(GYPA: AUC=0.861, P=0.012; ANK1: AUC=0.855, P=0.006; RHAG: AUC=0.854, P=0.009) was superior to that for 4, 8, 16, and 21 weeks(GYPA: AUC=0.646, 0.573, 0.691, and 0.617, respectively; ANK: AUC1=0.630, 0.658, 0.657, and 0.585, respectively; RHAG: AUC=0.592, 0.511, 0.515, and 0.536, respectively). Compared with the non-SONFH group, both SLC2A1 and STOM had the lowest expression levels in the peripheral blood of patients with the syndrome of liver and kidney deficiency(P<0.05). Compared with the normal control group, their expression levels were the lowest in the peripheral blood and affected femoral head tissue of SONFH rats modeled for 21 weeks(P<0.05, except STOM in the peripheral blood of rats). Moreover, the syndrome differentiation effectiveness of SLC2A1 in the rats modeled for 21 weeks(AUC=0.806, P=0.009) was superior to that for 4, 8, 12, and 16 weeks(AUC=0.520, 0.580, 0.741, 0.774, respectively), and STOM was meaningless in syndrome differentiation. In summary, the candidate marker gene for phlegm in SONFH is ELOVL6; the candidate marker genes for stasis are GYPA, RHAG, and ANK1; the candidate marker gene for deficiency is SLC2A1. The results help to reveal the biological connotations of phlegm, stasis, and deficiency in SONFH at the genetic level.

Endohedrally Functionalized Heteroleptic Coordination Cages for Phosphate Ester Binding.

Metallosupramolecular hosts of nanoscopic dimensions, which are able to serve as selective receptors and catalysts, are usually composed of only one type of organic ligand, restricting diversity in terms of cavity shape and functional group decoration. We report a series of heteroleptic [Pd2 A2 B2 ] coordination cages that self-assemble from a library of shape complementary bis-monodentate ligands in a non-statistical fashion. Ligands A feature an inward pointing NH function, able to engage in hydrogen bonding and amenable to being functionalized with amide and alkyl substituents. Ligands B comprise tricyclic aromatic backbones of different shape and electronic situation. The obtained heteroleptic coordination cages were investigated for their ability to bind phosphate diesters as guests. All-atom molecular dynamics (MD) simulations in explicit solvent were conducted to understand the mechanistic relationships behind the experimentally determined guest affinities.

Biosynthesis and Metabolism of Garlic Odor Compounds in Cultivated Chinese Chives (Allium tuberosum) and Wild Chinese Chives (Allium hookeri).

Chinese chives is a popular herb vegetable and medicine in Asian countries. Southwest China is one of the centers of origin, and the mountainous areas in this region are rich in wild germplasm. In this study, we collected four samples of germplasm from different altitudes: a land race of cultivated Chinese chives (Allium tuberosum), wide-leaf chives and extra-wide-leaf chives (Allium hookeri), and ovoid-leaf chives (Allium funckiaefolium). Leaf metabolites were detected and compared between A. tuberosum and A. hookeri. A total of 158 differentially accumulated metabolites (DAM) were identified by Gas Chromatography-Mass Spectrometry (GC-MS) and Liquid Chromatography-Mass Spectrometry (LC-MS), among which there was a wide range of garlic odor compounds, free amino acids, and sugars. A. hookeri contains a higher content of fructose, garlic odor compounds, and amino acids than A. tuberosum, which is supported by the higher expression level of biosynthetic genes revealed by transcriptome analysis. A. hookeri accumulates the same garlic odor compound precursors that A. tuberosum does (mainly methiin and alliin). We isolated full-length gene sequences of phytochelatin synthase (PCS), γ-glutamyltranspeptidases (GGT), flavin-containing monooxygenase (FMO), and alliinase (ALN). These sequences showed closer relations in phylogenetic analysis between A. hookeri and A. tuberosum (with sequence identities ranging from 86% to 90%) than with Allium cepa or Allium sativum (which had a lower sequence identity ranging from 76% to 88%). Among these assayed genes, ALN, the critical gene controlling the conversion of odorless precursors into odor compounds, was undetected in leaves, bulbs, and roots of A. tuberosum, which could account for its weaker garlic smell. Moreover, we identified a distinct FMO1 gene in extra-wide-leaf A. hookeri that is due to a CDS-deletion and frameshift mutation. These results above reveal the molecular and metabolomic basis of impressive strong odor in wild Chinese chives.