OBJECTIVETo explore the relation between gap junction and meridian phenomenon.

METHODSThe oxygen partial pressure in acupoints [see text for formula] and in their corresponding non-acupoints of the Bladder Meridian was observed with the needle-type tissue oxygen tension sensor in the gap junction blocking goats by 1-Heptanol injection and the Connexin 43 (Cx43) gene knockout mice.

RESULTS(1) The oxygen partial pressure in acupoints of Bladder Meridian on goats was higher than that in non-acupoints after 1-Heptanol injection with significant differences between them (both P < 0.01). (2) The oxygen partial pressure in acupoints of Bladder Meridian on goats increased significantly after injecting 1-Heptanol as compare with that either injecting normal saline or injecting nothing with significant differences between them (all P < 0.01). (3) The oxygen partial pressure in acupoints of the Bladder Meridian was significantly higher than that in the non-acupoint controls in Cx43 wild type (WT) mice (all P < 0.01). In Cx43 heterozygote (HT) mice, the oxygen partial pressure between acupoints and non-acupoint controls showed no significant differences (all P > 0.05). (4) In acupoints, the oxygen partial pressure in Cx43 WT mice was significantly higher than that in Cx43 HT mice (all P < 0.05), while in the corresponding non-acupoints, this difference had no statistically significant (all P > 0.05).

CONCLUSIONGap junction maybe the essential factor in signal transduction of acupuncture.

Acupuncture Points ; Animals ; Connexin 43 ; genetics ; metabolism ; Female ; Gap Junctions ; chemistry ; genetics ; metabolism ; Goats ; Male ; Meridians ; Mice ; Mice, Knockout ; Models, Animal ; Oxygen ; analysis ; metabolism ; Partial Pressure ; Urinary Bladder ; chemistry ; metabolism

OBJECTIVE: To analyze the molecular pathogenesis by analysis of phenotype and gene mutation in families with hereditary coagulation factor V (FⅤ) defect caused by complex heterozygous mutation. METHODS: Plasma pro-thrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), FⅤ procoagulant activity (FⅤ∶C), FⅤ antigen (FⅤ∶Ag), and other related coagulation indexes were detected in the proband and his family members (3 generations 10 people). Using DNA direct sequencing to analyze all exons, flanks, 5' and 3' untranslated regions of F5 genes and the corresponding mutation site regions of family members, the mutation site was confirmed by reverse sequencing．The conservation of mutant amino acids was analyzed by ClustalX-2.1-win software. The PROVEAN and MutationTaster online bioinformatics software were used to predict the effect of mutation on protein function. Protein model and amino acid interaction at mutation sites was analyzed by Swiss-pdbviewer software. RESULTS: The PT and APTT of the proband were significantly prolonged compared with healthy controls (34.2 vs 13.2 s and 119.3 vs 36.0 s), while FⅤ∶C and FⅤ∶Ag extremely reduced (3% and 6%). The PT and APTT of the second-born, the third son, daughter, and grandson of the proband were slightly prolonged, and the FⅤ∶C and FⅤ∶Ag decreased to varying degrees. The related coagulant parameters of other family members were within normal range. Genetic analysis revealed that the proband had a c.911G>A heterozygous missense mutation on the exon 6 lead to p.Gly276Glu, and a c.5343C>G heterozygous missense mutation on the exon 16 lead to p.Ser1781Arg of the proband. The second-born, the third son, and grandson of the proband carry p.Gly276Glu heterozygotes, and the daughter carries p.Ser1781Arg heterozygotes, while the other family members were wild-type. The results of conservative analysis indicated that p.Gly276 and p.Ser1781 were highly conserved in homologous species. The two bioinformatics software predicted the same results, PROVEAN (score -6.214 and -12.79) indicated that the compound heterozygous mutation was a harmful mutation; MutationTaster (score 0.976 and 0.999) suggested that these mutations might cause corresponding disease. p.Gly276Glu protein model analysis showed that, the Glu side chain was prolonged and the molecular weight became larger, which would increase the steric hindrance between it and the surrounding amino acids, affect the normal local folding of the FⅤ protein, and eventually lead to the decrease of protein activity and content. This paper can not provide analysis of the spatial structure of p.Ser1781Arg mutant protein because of the lack of X ray 3 D structure file of FⅤ exon 16. CONCLUSION The new compound heterozygous mutations (p.Gly276Glu and p.Ser1781Arg) identified in this study are the main reasons for the decrease in the FⅤ level of the family, among which p.Ser1781Arg is rarely reported at home and abroad.
Factor V/genetics* ; Family ; Genotype ; Heterozygote ; Humans ; Mutation ; Pedigree ; Phenotype

OBJECTIVETo investigate the effect and mechanism of miR-486 on glycometabolism of hematopoietic cells.

METHODSqRT-PCR was applied to detect the expression of miR-486 or Sirt1 on TF-1 cells under hypoxia. Lentivirus was used to mediate the overexpression or inhibition of miR-486 on TF-1 cells and qRT-PCR was used to detect the expressions of Sirt1, glucose transporter 1(Glut1) and glucose transporter 4(Glut4). Lentivirus-mediated Sirt1-shRNA transduction was used to knockdown Sirt1 expression which was detected by qRT-PCR and Western blot. The expressions of Glut1 and Glut4 were determined by qRT-PCR.

RESULTSHypoxia promoted the expression of miR-486 and inhibited the expression of Sirt1. MiR-486 overexpression could inhibit the expression of Sirt1 and promote the expressions of Glut1 and Glut4, whereas miR-486 silencing upregulated the sirt1 expression and inhibited the expressions of Glut1 and Glut4. And inhibition of Sirt1 expression increased the expressions of Glut1 and Glut4.

CONCLUSIONMiR-486 can regulate the glycometabolism of hematopoietic cells by targeting Sirt1.