Cbfa1 is a critical regulator of cell differentiation expressed only in the osteochondrogenic lineage. To define the molecular basis of this cell-specific expression we analyzed the murine Cbfa1 promoter. Here we show that the first 976 bp of this promoter are specifically active in osteoblastic cells. Within this region DNase I footprinting delineated a 40-bp area (CE1) protected differently by nuclear extracts from osteoblastic cells and from non-osteoblastic cells. When multimerized, CE1 conferred an osteoblast-specific activity to a heterologous promoter in DNA transfection experiments; this enhancing ability was conserved between mouse, rat, and human CE1 present in the respective Cbfa1 promoters. CE1 site-specific mutagenesis determined that it binds NF1-and AP1-like activities. Further analyses revealed that the NF1 site acts as a repressor in nonosteoblastic cells due to the binding of NF1-A, a NF1 isoform not expressed in osteoblastic cells. In contrast, the AP1 site mediates an osteoblast-specific activation caused by the preferential binding of FosB to CE1 in osteoblastic cells. In summary, this study identified an osteoblast-specific enhancer in the Cbfa1 promoter whose activity is achieved by the combination of an inhibitory and an activatory mechanism.

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Continuous 24-hr intragastric pH-metry was prospectively performed in 801 subjects with different clinical conditions using two pH electrodes placed closely adjacent. The aim was to assess the in situ repeatability of the test and to verify whether the removal of artifacts, interference, and noise usually superimposed onto the fundamental signal recorded by the measuring apparatus improves the clinical usefulness of experimental data. The following debugging/filtering procedure was used: first, pH recordings of each channel were amended separately from artifacts, then they underwent 7 min windowed median interference debugging, and finally Wiener noise filtering was applied. Afterwards, the 24-hr mean pH profile was obtained in each subject by averaging the pH tracings of the two channels every minute (1440 data points/24 hr). The efficiency of this procedure was assessed at each step by evaluating the difference among groups using the O'Brien test, a distribution-free nonparametric method well-suited for evaluating differences among groups allocated onto a two-way layout. The differences among groups calculated from raw pH data of the single channels can be very misleading, in that it is possible to find that they are significant on one channel and not significant on the other channel. Conversely, the significance of the differences among groups increases progressively at each step of the above debugging/filtering procedure applied to raw pH profiles of each channel. Seven minutes was shown to be the most suitable time lag for windowed median removal of interference.(ABSTRACT TRUNCATED AT 250 WORDS)