Olig1 and ID4 interactions in living cells visualized by bimolecular fluorescence complementation technique.

Olig1, a member of class B basic-helix-loop-helix (bHLH), plays key roles in early oligodendrocyte specification. Inhibitors of DNA binding (Id) is another sub-class of HLH proteins, act as dominant-negative regulators of bHLH proteins, which can form heterodimers with class A or B bHLH proteins, but lack the critical basic DNA binding domain. Id4 was recently found to interact with olig1 and inhibit oligodendrocyte differentiation. However, there still no direct evidence to reveal the spatial and temporal interaction of olig1 and ID4 in living cells. In this study, we performed bimolecular fluorescence complementation (BiFC) analysis to further characterize the distinct subcellular localization of olig1, ID4 and their dimer in living SW1116 cells. To examine the subcellular localization of olig1 and ID4 by themselves, the olig1-EGFP or ID4-DsRed2 fusion proteins were also expressed in SW1116 cells, respectively. As predicted, the olig1-EGFP fusion proteins were located in the nucleus, and ID4-DsRed2 fusion proteins were located in the cytoplasm. When olig1-EGFP and ID4-DsRed2 fusion proteins were co-expressed, the green and red signals were co-located in the cytoplasm. Using BiFC, the strong BiFC signals could be detected in pBiFC-olig1VN173 and pBiFC-ID4VC155 co-transfected cells and the fluorescence signal was located in the cytoplasm. These results collectively confirmed that olig1 and ID4 could interact and form dimer in living cells, and ID4 could block the transport of olig1 from cytoplasm to nucleus.

BMP signaling mediates astrocyte differentiation of oligodendrocyte progenitor cells.

Oligodendrocyte precursor cells (OPCs) can differentiate into oligodendrocytes or astrocytes, depending on cellular microenvironments. OPCs, cultured in medium supplemented with 10% (v/v) fetal bovine serum (FBS), give rise to type II astrocytes that express glial fibrillary acidic protein and a cell surface ganglioside that is recognized by A2B5 monoclonal antibody. However, the factors in FBS that direct the astrocyte differentiation are not determined. Moreover, bone morphogenetic proteins (BMPs) have been reported to be involved in astrocyte differentiation of neural progenitor cells. We therefore examined whether BMPs are responsible for the serum-mediated astrocyte differentiation from OPCs. OPCs were isolated from the spinal cords of Wistar rat embryos (at day 14) using the A2B5 antibody. We measured the concentrations of BMP-2 and BMP-4 in FBS and rat and human sera and the expression of mRNAs for three types of BMP receptors (BMPRIa, Ib and II) in OPCs by RT-PCR. The serum samples of the three species contained BMP-2 and BMP-4, as judged by ELISA with each monoclonal antibody, and the BMP receptor mRNAs are expressed in OPCs. When OPCs were cultured in the medium containing 10% FBS, cells (more than 95%) differentiated into type II astrocytes. However, when OPCs were pretreated with noggin, a soluble antagonist of BMP action, the degree of astrocyte differentiation was markedly decreased from 95.39 to 38.36%. Taken together, these results suggest that BMP signaling may be responsible for the serum-mediated astrocyte differentiation of OPCs. Our findings provide new insights into the molecular basis of differentiation of OPCs.

[A multi-center, randomized, double-blind, placebo-controlled trial of glyceryl trinitrate ointment in the treatment of anal fissure].

OBJECTIVE: To evaluate the clinical efficacy and safety of glyceryl trinitrate (GTN) ointment in the treatment of anal fissure. METHODS: In this multi-center, randomized, double-blind and placebo-controlled trial, 240 chronic anal fissure patients from 7 clinical centers were randomized to receive eight-week treatment with GTN ointment (treatment group) or vaseline ointment (control group) respectively. Healing rate, visual analogue score (VAS), maximum anal resting pressure (MARP) and adverse reactions were recorded and compared. RESULTS: A total of 221 patients (92.1%) finished the trial, including 114 patients in treatment group (95.0%, 114/120) and 107 in control group (89.2%, 107/120). At the endpoint of treatment (56 d), 90 patients in treatment group (78.9%, 90/114) healed completely compared to 31 patients in control group (29.0%, 31/107), and decrease rates of VAS in the two groups were (94.8±15.7)% and (61.2±35.7)% respectively, both differences were statistically significant (P<0.01). MARP after first administration was (20.2±18.5) mm Hg in treatment group (n=12) and (7.1±14.7) mm Hg in control group (n=6), which was not significantly different (P=0.152). Adverse reaction incidence was higher in treatment group (42.1% vs. 9.3%, P<0.05), while these adverse reactions were mainly headache and fullness in head, which were self-limiting. CONCLUSION: GTN ointment can effectively promote healing and relieve pain in anal fissure with safety and tolerance.