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Ligands of the transforming growth factor-beta (TGF-beta) superfamily are involved in numerous developmental and disease processes. TGF-beta, activins, and nodal ligands operate through the highly homologous Smad2 and Smad3 intracellular mediators. Smad2 mutants exhibit early embryonic lethality, while Smad3 mutants are viable, but show a plethora of postnatal phenotypes, including immune dysfunction and skeletal abnormalities. Previously, we have shown that the Smad2 and Smad3 genes function cooperatively during liver morphogenesis. Here we show that Smad2 and Smad3 are required at a full dosage for normal embryonic development. Animals lacking one allele of each gene exhibit a variably penetrant phenotype in which structures in the anterior and ventral midline are reduced or lost; additionally, we demonstrate that this craniofacial defect and the previously reported hepatic phenotypes are both due to defects in the definitive endoderm. A reduction of endodermal gene expression as well as a failure to displace the visceral endoderm occurs despite the formation of a normal foregut pocket. This precedes any defects in anterior patterning and likely causes the abnormalities observed in craniofacial and midline development, as well as hepatogenesis. Keywords: Endoderm; Smad; TGF-beta; Holoprosencephaly; Hex; Foxa2; Craniofacial; Liver

Background: A spectrophotometric serum lithium assay and relevant reaction equilibria used in the assay development are described. Methods: The dye, 7-nitro-2,12-dicarboxyl-16,17-dihydro-5H,15H-dibenzo[b,i] [1,11,4,5,7,8]dioxatetraazacyclotetradecine reacts rapidly with Li+ in alkaline water/dimethylsulfoxide with an absorbance change. Sodium binds weakly, thus presenting potential inaccuracy from varying endogenous sodium. To minimize this problem, excess sodium ion was added from reagents. This diminishes the signal from lithium binding, thus lowering sensitivity, requiring selection of reagent sodium concentration for proper balance of performance characteristics. Absorptivities and binding constants were determined. Description of competing equilibria by an explicit cubic equation was used to predict sensitivity and effects of varying endogenous Na+ concentration as functions of added NaCl. Results: Concentration-absorbance data were satisfactorily predicted by the cubic equation, equilibrium constants, and absorptivities. Li+ binds 300 times more tightly than Na+. Interference was minimized, while sufficient sensitivity for needed precision was retained. A full evaluation is summarized separately in this issue. Conclusions: Explicit description of the competitive binding was successfully used to develop an automated liquid reagent assay for Li+ with acceptable accuracy and precision. [Copyright &y& Elsevier]

Summary: The evolutionarily ancient arm of the E2f family of transcription factors consisting of the two atypical members E2f7 and E2f8 is essential for murine embryonic development. However, the critical tissues, cellular processes, and molecular pathways regulated by these two factors remain unknown. Using a series of fetal and placental lineage-specific cre mice, we show that E2F7/E2F8 functions in extraembryonic trophoblast lineages are both necessary and sufficient to carry fetuses to term. Expression profiling and biochemical approaches exposed the canonical E2F3a activator as a key family member that antagonizes E2F7/E2F8 functions. Remarkably, the concomitant loss of E2f3a normalized placental gene expression programs, corrected placental defects, and fostered the survival of E2f7/E2f8-deficient embryos to birth. In summary, we identified a placental transcriptional network tightly coordinated by activation and repression through two distinct arms of the E2F family that is essential for extraembryonic cell proliferation, placental development, and fetal viability. [Copyright &y& Elsevier]

Background: Therapeutic monitoring of lithium is important because of its narrow therapeutic range and therapeutic index, low protein binding and single route of elimination. We characterized a new photometric method that avoids the specialized requirements of ion-specific electrode (ISE), atomic absorption and flame emission methods. Methods: Minimum detectable concentration (MDC), linearity and calibration drift over 65 days were determined. Within-run, between-run and total imprecision were assessed over 20 days in accordance with NCCLS EP5. Interference studies were conducted for 46 endogenous and exogenous compounds. Two production lots of the new photometric method (LI) were compared on the Dimension® RxL system and two ISE methods [Ciba Corning (n=124) and DuPont (n=131)], an established photometric method (Vitros® 950 system; n=63) and atomic absorption (Thermo-Jerell Ash; n=63). Results: The MDC was 0.04 mmol/l. Linearity was demonstrated from 0.12 to 5.8 mmol/l by the regression equation: observed=(1.01×expected)−0.0005 mmol/l, Sy/x=0.03 mmol/l, r=0.999. Drift for the lithium calibrators over the 65-day study period was <5%, except for the lowest calibrator, which showed 0.04 mmol/l drift. None of the 46 potential interfering substances showed greater than a 6.5% difference between control and test solutions. ISE method comparisons showed the following: LI=(1.08×Ciba Corning ISE)−0.15 mmol/l, Sy/x=0.05, r=0.999, and LI=(1.03×DuPont ISE)+0.00 mmol/l, Sy/x=0.06 mmol/l, r=0.999. Comparison of the LI method with the atomic absorption and Vitros® system showed proportionality error <10%. Bias between the LI method and atomic absorption was 7%, substantially less than that documented in proficiency surveys for the Vitros® and other systems. No lot-to-lot or site-to-site differences were observed. Conclusion: This new photometric method is an attractive alternative for Li measurement and is adaptable to instruments having spectrophotometric capability. [Copyright &y& Elsevier]