Bone morphogenetic proteins, extracellular matrix, and mitogen-activated protein kinase signaling pathways are required for osteoblast-specific gene expression and differentiation in MC3T3-E1 cells.

Osteoblasts secrete a complex extracellular matrix (ECM) containing collagenous and noncollagenous proteins, bone morphogenetic proteins (BMPs), and growth factors. Osteoblast-specific gene expression requires ascorbic acid (AA)-dependent assembly of a collagenous ECM. Matrix responsiveness requires an alpha2beta1 integrin-collagen interaction and mitogen-activated protein kinase (MAPK) activity, which phosphorylates and activates the osteoblast-specific transcription factor Cbfa1. This study examines interactions between this integrin/MAPK-mediated pathway and signals initiated by BMPs contained in the osteoblast matrix. MC3T3-E1 cells were shown to constitutively express BMP-2, BMP-4, and BMP-7. Noggin, a specific BMP inhibitor, reversibly blocked AA-induced gene expression, indicating that BMP production by MC3T3-E1 cells was necessary for differentiation. The ability of exogenously added BMP-2, BMP-4, or BMP-7 to stimulate osteocalcin (OCN) and bone sialoprotein (BSP) mRNAs or OCN promoter activity was synergistically increased in cells that were actively synthesizing an ECM (i.e., were grown in the presence of AA). A minimum of 4 days of ECM accumulation was required for this synergistic response to be observed. Neither BMP-7, AA, nor a combination of these two treatments had major effects on Cbfa1 messenger RNA (mRNA) or protein levels, as would be expected if regulation was mainly at the posttranscriptional level. U0126, a specific inhibitor of MAPK/extracellular signal-regulated kinase (MEK), blocked AA- or BMP-7/AA-dependent gene expression in a time- and dose-dependent manner that was closely correlated with inhibition of extracellular signal-regulated kinase (ERK) phosphorylation. This work establishes that autocrine BMP production as well as integrin-mediated cell-collagen interactions are both required for osteoblast differentiation, and both these pathways require MAP kinase activity.

Utility of ultrasound-guided central venous cannulation in pediatric surgical patients: a clinical series.

BACKGROUND: Central venous cannulation can be particularly difficult in pediatric patients. Central line placement is associated with many well-known complications. While ultrasound-guided techniques are well established, the majority of central venous catheters are placed using landmark guidance. This retrospective study compares the safety and efficacy of ultrasound guidance vs landmark guidance in central venous cannulation of pediatric cardiac surgery patients. METHODS: The medical records of 149 pediatric patients undergoing cardiac surgery over 3-year period were reviewed. Patients were classified into two cohorts based on whether central venous cannulation of the internal jugular vein was performed by ultrasound or landmark guidance. Overall success and traumatic complication rates were compared between the two groups. Additionally, comparisons between the groups were made to determine if patient size or age affected the success rate of either approach in different manner. RESULTS: Patients in the ultrasound-guided (n = 47) and the landmark-guided (n = 102) groups were similar with respect to age, weight, and surgical procedure for which central venous access was indicated. The overall success rate for cannulation of the internal jugular vein was 91.5% in the ultrasound-guided group and 72.5% in the landmark-guided group (P = 0.010). But in the subgroup of children under 1 year of age, success rate was 77.8% in ultrasound group and 60.9% in landmark group (P = 0.44); in children under 10 kg in weight, success rate was 80% in ultrasound group and 56.7% in landmark group (P = 0.19). There were no significant differences in the rate of traumatic complications between the two methods. CONCLUSIONS: The overall success of internal jugular vein cannulation for pediatric cardiac surgery is significantly improved with the use of ultrasound guidance, without a significant difference in traumatic complications. However, mostly children above 1 year of age or 10 kg of weight experience advantages of ultrasound technique.

Caudal anesthesia in pediatric cardiac surgery: does it affect outcome?

OBJECTIVE: The purpose of this study was to examine the influence of caudal anesthesia on outcomes (pediatric intensive care unit [PICU] length of stay, hospital length of stay, ventilatory time, early extubation rate) in pediatric patients undergoing congenital heart disease repair requiring cardiopulmonary bypass (CPB). DESIGN: Retrospective. SETTING: University teaching hospital. PARTICIPANTS: Pediatric patients undergoing surgery to treat congenital heart disease between 1999 and 2002. INTERVENTION: None. MEASUREMENTS AND MAIN RESULTS: Thirty-four patients with atrial septal defect (ASD), 37 with ventricular septal defect, and 46 with tetralogy of Fallot (TOF) were included in the analysis. No differences were found in preoperative and intraoperative data between caudal and noncaudal group for each disorder. There was no difference between caudal and noncaudal groups in PICU and hospital stay. A statistically significant difference was found in the postoperative ventilatory time in patients with ASD and TOF between caudal and noncaudal groups. The early extubation rate was higher in the TOF caudal group compared with the noncaudal group. CONCLUSIONS: This retrospective study demonstrated that postinduction placement of caudal anesthesia does not affect PICU or hospital length of stay. A well-controlled prospective study is needed to confirm these findings.

Multiple signaling pathways converge on the Cbfa1/Runx2 transcription factor to regulate osteoblast differentiation.

The Cbfa1/Runx2 transcription factor is essential for osteoblast differentiation. However, levels of Runx2 are often not well correlated with its transcriptional activity suggesting that this factor must be activated either by covalent modification or through interactions with other nuclear components. Runx2 is phosphorylated and activated by the mitogen-activated protein kinase (MAPK) pathway. This pathway is stimulated in at least two ways: by binding of type I collagen to alpha2beta1 integrins on the osteoblast surface and by treatment of cells with the osteogenic growth factor, FGF2. Protein kinase A (PKA) also may phosphorylate/activate Runx2 under certain conditions. Runx2 activity also is enhanced by factors known to stimulate specific signal transduction pathways such as PTH/PTHrP (signals through PKA and PKC pathways) and BMPs (Signal through Smad proteins). Interactions with Runx2 are complex involving both binding of distinct components such as AP-1 factors and Smads to separate sites on DNA, direct interactions between Runx2 and AP-1/Smad factors and MAPK or PKA-dependent Runx2 phosphorylation. These findings suggest that Runx2 plays a central role in coordinating multiple signals involved in osteoblast differentiation.