A Late Role for bmp2b in the Morphogenesis of Semicircular Canal Ducts in the Zebrafish Inner Ear

Background

The Bone Morphogenetic Protein (BMP) genes bmp2 and bmp4 are expressed in highly conserved patterns in the developing vertebrate inner ear. It has, however, proved difficult to elucidate the function of BMPs during ear development as mutations in these genes cause early embryonic lethality. Previous studies using conditional approaches in mouse and chicken have shown that Bmp4 has a role in semicircular canal and crista development, but there is currently no direct evidence for the role of Bmp2 in the developing inner ear.

Methodology/Principal Findings

We have used an RNA rescue strategy to test the role of bmp2b in the zebrafish inner ear directly. Injection of bmp2b or smad5 mRNA into homozygous mutant swirl (bmp2b^−/−) embryos rescues the early patterning defects in these mutants and the fish survive to adulthood. As injected RNA will only last, at most, for the first few days of embryogenesis, all later development occurs in the absence of bmp2b function. Although rescued swirl adult fish are viable, they have balance defects suggestive of vestibular dysfunction. Analysis of the inner ears of these fish reveals a total absence of semicircular canal ducts, structures involved in the detection of angular motion. All other regions of the ear, including the ampullae and cristae, are present and appear normal. Early stages of otic development in rescued swirl embryos are also normal.

Conclusions/Significance

Our findings demonstrate a critical late role for bmp2b in the morphogenesis of semicircular canals in the zebrafish inner ear. This is the first demonstration of a developmental role for any gene during post-embryonic stages of otic morphogenesis in the zebrafish. Despite differences in the early stages of semicircular canal formation between zebrafish and amniotes, the role of Bmp2 in semicircular canal duct outgrowth is likely to be conserved between different vertebrate species.     

Are Cytochrome P450 CYP2C8 and CYP2C9 Polymorphisms Associated with Ibuprofen Response in Very Preterm Infants?

Background

Patent ductus arteriosus (PDA) in extremely preterm infants remains a challenging condition with conflicting treatment strategies. Ibuprofen is currently used to treat PDA with ductal closure failure rate up to 40%. We test the hypothesis that cytochrome P450 CYP2C8/2C9 polymorphisms may predict ibuprofen response.

Methodology/Principal Findings

We studied extremely preterm neonates with haemodynamically significant PDA and treated with ibuprofen. One or two variant CYP2C8 and/or 2C9 alleles were found in 17% of the population, most of them were from Caucasian ethnicity (67–74%). Response to ibuprofen and clinical course of infants carrying variants CYP2C8 and CYP2C9 were similar. Comparing infants with wild type or variant CYP2C8 and CYP2C9 genotypes, response rate to ibuprofen was significantly higher in wild type than in mutated carriers in univariate analysis (73% versus 52%, p = 0.04). Comparing responders (ductus closure; n = 75) and non-responders (surgical ligation; n = 36), the only two factors significantly associated with the response to ibuprofen using multivariate analysis were higher gestational age and non Caucasian ethnicity but not CYP2C polymorphism.

Conclusions

CYP2C polymorphism was not associated with PDA response to ibuprofen and this factor appears not appropriate to optimize the ductal closure rate by modulating ibuprofen dosing strategy. This study points out the role for ethnicity in the interindividual variability of response to ibuprofen in extremely preterm infants.     

Bmp7 functions via a polarity mechanism to promote cloacal septation

Background

During normal development in human and other placental mammals, the embryonic cloacal cavity separates along the axial longitudinal plane to give rise to the urethral system, ventrally, and the rectum, dorsally. Defects in cloacal development are very common and present clinically as a rectourethral fistula in about 1 in 5,000 live human births. Yet, the cellular mechanisms of cloacal septation remain poorly understood.

Methodology/Principal Findings

We previously detected Bone morphogenetic protein 7 (Bmp7) expression in the urorectal mesenchyme (URM), and have shown that loss of Bmp7 function results in the arrest of cloacal septation. Here, we present evidence that cloacal partitioning is driven by Bmp7 signaling in the cloacal endoderm. We performed TUNEL and immunofluorescent analysis on cloacal sections from Bmp7 null and control littermate embryos. We found that loss of Bmp7 results in a dramatic decrease in the endoderm survival and a delay in differentiation. We used immunological methods to show that Bmp7 functions by activating the c-Jun N-terminal kinase (JNK) pathway. We carried out confocal and 3D imaging analysis of mitotic chromosome bundles to show that during normal septation cells in the cloacal endoderm divide predominantly in the apical-basal direction. Loss of Bmp7/JNK signaling results in randomization of mitotic angles in the cloacal endoderm. We also conducted immunohistochemical analysis of human fetal sections to show that BMP/phospho-SMAD and JNK pathways function in the human cloacal region similar as in the mouse.

Conclusion/Significance

Our results strongly indicate that Bmp7/JNK signaling regulates remodeling of the cloacal endoderm resulting in a topological separation of the urinary and digestive systems. Our study points to the importance of Bmp and JNK signaling in cloacal development and rectourethral malformations.     

Comparison of Oral Paracetamol versus Ibuprofen in Premature Infants with Patent Ductus Arteriosus: A Randomized Controlled Trial

Trial Design

Oral ibuprofen has demonstrated good effects on symptomatic patent ductus arteriosus (PDA) but with many contraindications and potential side-effects. In the past two years, oral paracetamol administration to several preterm infants with PDA has been reported. Here, a randomized, non-blinded, parallel-controlled and non-inferiority trial was designed to evaluate the efficacy and safety profiles of oral paracetamol to those of standard ibuprofen for PDA closure in premature infants.

Methods

One hundred and sixty infants (gestational age ≤34 weeks) with echocardiographically confirmed PDA were randomly assigned to receive either oral paracetamol (n = 80) or ibuprofen (n = 80). After the initial treatment course in both groups, the need for a second course was determined by echocardiographic evaluation. The main outcome was rate of ductal closure, and secondary outcomes were adverse effects and complications.

Result

The ductus was closed in 65 (81.2%) infants of the paracetamol group compared with 63 (78.8%) of the ibuprofen group. The 95% confidence interval of the difference between these groups was [−0.080,0.128], demonstrating that the effectiveness of paracetamol treatment was not inferior to that of ibuprofen. In fact, the incidence of hyperbilirubinemia or gastrointestinal bleeding in the paracetamol group was significantly lower than that of the ibuprofen group. No significant differences in other clinical side effects or complications were noted.

Conclusion

This comparison of drug efficacy and safety profiles in premature infants with PDA revealed that oral paracetamol was comparable to ibuprofen in terms of the rate of ductal closure and even showed a decreased risk of hyperbilirubinemia or gastrointestinal bleeding. Therefore, paracetamol may be accepted as a first-line drug treatment for PDA in preterm infants.

Trial Registration

ChiCTR.org ChiCTR-TRC-12002177     

Twist controls skeletal development and dorsoventral patterning by regulating runx2 in zebrafish

Background

Twist1a and twist1b are the principal components of twists that negatively regulate a number of cellular signaling events. Expression of runx2 and downstream targets is essential for skeletal development and ventral organizer formation and specification in early vertebrate embryos, but what controls ventral activity of maternal runx2 and how twists function in zebrafish embryogenesis still remain unclear.

Methodology/Principal Findings

By studying the loss of twist induced by injection of morpholino-oligonucleotide in zebrafish, we found that twist1a and twist1b, but not twist2 or twist3, were required for proper skeletal development and dorsoventral patterning in early embryos. Overexpression of twist1a or twist1b following mRNA injection resulted in deteriorated skeletal development and formation of typical dorsalized embryos, whereas knockdown of twist1a and twist1b led to the formation of abnormal embryos with enhanced skeletal formation and typical ventralized patterning. Overexpression of twist1a or twist1b decreased the expression of runx2b, whereas twist1a and twist1b knockdown increased runx2b expression. We have further demonstrated that phenotypes induced by twist1a and twist1b knockdown were rescued by runx2b knockdown.

Conclusions/Significance

Together, these results suggest that twist1a and twist1b control skeletal development and dorsoventral patterning by regulating runx2b in zebrafish and provide potential targets for the treatment of diseases or syndromes associated with decreased skeletal development.     

Left-right function of dmrt2 genes is not conserved between zebrafish and mouse

Background

Members of the Dmrt family, generally associated with sex determination, were shown to be involved in several other functions during embryonic development. Dmrt2 has been studied in the context of zebrafish development where, due to a duplication event, two paralog genes dmrt2a and dmrt2b are present. Both zebrafish dmrt2a/terra and dmrt2b are important to regulate left-right patterning in the lateral plate mesoderm. In addition, dmrt2a/terra is necessary for symmetric somite formation while dmrt2b regulates somite differentiation impacting on slow muscle development. One dmrt2 gene is also expressed in the mouse embryo, where it is necessary for somite differentiation but with an impact on axial skeleton development. However, nothing was known about its role during left-right patterning in the lateral plate mesoderm or in the symmetric synchronization of somite formation.

Methodology/Principal Findings

Using a dmrt2 mutant mouse line, we show that this gene is not involved in symmetric somite formation and does not regulate the laterality pathway that controls left-right asymmetric organ positioning. We reveal that dmrt2a/terra is present in the zebrafish laterality organ, the Kupffer''s vesicle, while its homologue is excluded from the mouse equivalent structure, the node. On the basis of evolutionary sub-functionalization and neo-functionalization theories we discuss this absence of functional conservation.

Conclusions/Significance

Our results show that the role of dmrt2 gene is not conserved during zebrafish and mouse embryonic development.     

Expression and Functional Characterization of Smyd1a in Myofibril Organization of Skeletal Muscles

Background

Smyd1, the founding member of the Smyd family including Smyd-1, 2, 3, 4 and 5, is a SET and MYND domain containing protein that plays a key role in myofibril assembly in skeletal and cardiac muscles. Bioinformatic analysis revealed that zebrafish genome contains two highly related smyd1 genes, smyd1a and smyd1b. Although Smyd1b function is well characterized in skeletal and cardiac muscles, the function of Smyd1a is, however, unknown.

Methodology/Principal Findings

To investigate the function of Smyd1a in muscle development, we isolated smyd1a from zebrafish, and characterized its expression and function during muscle development via gene knockdown and transgenic expression approaches. The results showed that smyd1a was strongly expressed in skeletal muscles of zebrafish embryos. Functional analysis revealed that knockdown of smyd1a alone had no significant effect on myofibril assembly in zebrafish skeletal muscles. However, knockdown of smyd1a and smyd1b together resulted in a complete disruption of myofibril organization in skeletal muscles, a phenotype stronger than knockdown of smyd1a or smyd1b alone. Moreover, ectopic expression of zebrafish smyd1a or mouse Smyd1 transgene could rescue the myofibril defects from the smyd1b knockdown in zebrafish embryos.

Conclusion/Significance

Collectively, these data indicate that Smyd1a and Smyd1b share similar biological activity in myofibril assembly in zebrafish embryos. However, Smyd1b appears to play a major role in this process.     

A comparison of postoperative outcomes with PDA ligation in the OR versus the NICU: a retrospective cohort study on the risks of transport

Background

Although patent ductus arteriosus (PDA) ligations in the Neonatal Intensive Care Unit (NICU) have been an accepted practice, many are still performed in the Operating Room (OR). Whether avoiding transport leads to improved perioperative outcomes is unclear. Here we aimed to determine whether PDA ligations in the NICU corresponded to higher risk of surgical site infection or mortality and if transport was associated with worsened perioperative outcomes.

Methods

We performed a retrospective cohort study of NICU patients, ≤37?weeks post-menstrual age, undergoing surgical PDA ligation in the NICU or OR. We excluded any infants undergoing device PDA closure. We measured the incidence of perioperative hypothermia, cardiac arrest, decreases in SpO2, hemodynamic instability and postoperative surgical site infection, sepsis and mortality.

Results

Data was collected on 189 infants (100 OR, 89 NICU). After controlling for number of preoperative comorbidities, weight at time of procedure, procedure location and hospital in the mixed-effect model, no significant difference in mortality or sepsis was found (odds ratio 0.31, 95%CI 0.07, 1.30; p?=?0.107, and odds ratio 0.40; 95%CI 0.14, 1.09; p?=?0.072, respectively). There was an increased incidence of hemodynamic instability on transport postoperatively in the OR group (12.4% vs 2%, odds ratio 6.93; 95% CI 1.48, 35.52; p?=?0.014).

Conclusion

PDA ligations in the NICU were not associated with higher incidences of surgical site infection or mortality. There was an increased incidence of hemodynamic instability in the OR group on transport back to the NICU. Larger multicenter studies following long-term outcomes are needed to evaluate the safety of performing all PDA ligations in the NICU.

Keywords

Patent ductus arteriosus, Newborn infant, Neonatal intensive care unit, Surgical wound infection, Postoperative period, Hemodynamics

     

Overlapping mouse subcongenic strains successfully separate two linked body fat QTL on distal MMU 2

Background

Mouse chromosome 2 is linked to growth and body fat phenotypes in many mouse crosses. With the goal to identify the underlying genes regulating growth and body fat on mouse chromosome 2, we developed five overlapping subcongenic strains that contained CAST/EiJ donor regions in a C57BL/6J^hg/hg background (hg is a spontaneous deletion of 500 Kb on mouse chromosome 10). To fine map QTL on distal mouse chromosome 2 a total of 1,712 F2 mice from the five subcongenic strains, plus 278 F2 mice from the HG2D founder congenic strain were phenotyped and analyzed. Interval mapping (IM) and composite IM (CIM) were performed on body weight and body fat traits on a combination of SNP and microsatellite markers, which generated a high-density genotyping panel.

Results

Phenotypic analysis and interval mapping of total fat mass identified two QTL on distal mouse chromosome 2. One QTL between 150 and 161 Mb, Fatq2a, and the second between 173.3 and 175.6 Mb, Fatq2b. The two QTL reside in different congenic strains with significant total fat differences between homozygous cast/cast and b6/b6 littermates. Both of these QTL were previously identified only as a single QTL affecting body fat, Fatq2. Furthermore, through a novel approach referred here as replicated CIM, Fatq2b was mapped to the Gnas imprinted locus.

Conclusions

The integration of subcongenic strains, high-density genotyping, and CIM succesfully partitioned two previously linked QTL 20 Mb apart, and the strongest QTL, Fatq2b, was fine mapped to a ~2.3 Mb region interval encompassing the Gnas imprinted locus.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-014-1191-8) contains supplementary material, which is available to authorized users.     

AtPV42a and AtPV42b redundantly regulate reproductive development in Arabidopsis thaliana

Background

The conserved SNF1/AMPK/SnRK1 complexes are global regulators of metabolic responses in eukaryotes and play a key role in the control of energy balance. Although α-type subunits of the SnRK1 complex have been characterized in several plant species, the biological function of β-type and γ-type subunits remains largely unknown. Here, we characterized AtPV42a and AtPV42b, the two homologous genes in Arabidopsis, which encode cystathionine-β-synthase (CBS) domain-containing proteins that belong to the PV42 class of γ-type subunits of the plant SnRK1 complexes.

Methodology/Principal Findings

Real-time polymerase chain reaction was performed to examine the expression of AtPV42a and AtPV42b in various tissues. Transgenic plants that expressed artificial microRNAs targeting these two genes were created. Reproductive organ development and fertilization in these plants were examined by various approaches, including histological analysis, scanning electron microscopy, transmission electron microscopy, and phenotypic analyses of reciprocal crosses between wild-type and transgenic plants. We found that AtPV42a and AtPV42b were expressed in various tissues during different developmental stages. Transgenic plants where AtPV42a and AtPV42b were simultaneously silenced developed shorter siliques and reduced seed sets. Such low fertility phenotype resulted from deregulation of late stamen development and impairment of pollen tube attraction conferred by the female gametophyte.

Conclusions

Our results demonstrate that AtPV42a and AtPV42b play redundant roles in regulating male gametogenesis and pollen tube guidance, indicating that the Arabidopsis SnRK1 complexes might be involved in the control of reproductive development.     

Ectodomains of the LDL Receptor-Related Proteins LRP1b and LRP4 Have Anchorage Independent Functions In Vivo

Background

The low-density lipoprotein (LDL) receptor gene family is a highly conserved group of membrane receptors with diverse functions in developmental processes, lipoprotein trafficking, and cell signaling. The low-density lipoprotein (LDL) receptor-related protein 1b (LRP1B) was reported to be deleted in several types of human malignancies, including non-small cell lung cancer. Our group has previously reported that a distal extracellular truncation of murine Lrp1b that is predicted to secrete the entire intact extracellular domain (ECD) is fully viable with no apparent phenotype.

Methods and Principal Findings

Here, we have used a gene targeting approach to create two mouse lines carrying internally rearranged exons of Lrp1b that are predicted to truncate the protein closer to the N-terminus and to prevent normal trafficking through the secretary pathway. Both mutations result in early embryonic lethality, but, as expected from the restricted expression pattern of LRP1b in vivo, loss of Lrp1b does not cause cellular lethality as homozygous Lrp1b-deficient blastocysts can be propagated normally in culture. This is similar to findings for another LDL receptor family member, Lrp4. We provide in vitro evidence that Lrp4 undergoes regulated intramembraneous processing through metalloproteases and γ-secretase cleavage. We further demonstrate negative regulation of the Wnt signaling pathway by the soluble extracellular domain.

Conclusions and Significance

Our results underline a crucial role for Lrp1b in development. The expression in mice of truncated alleles of Lrp1b and Lrp4 with deletions of the transmembrane and intracellular domains leads to release of the extracellular domain into the extracellular space, which is sufficient to confer viability. In contrast, null mutations are embryonically (Lrp1b) or perinatally (Lrp4) lethal. These findings suggest that the extracellular domains of both proteins may function as a scavenger for signaling ligands or signal modulators in the extracellular space, thereby preserving signaling thresholds that are critical for embryonic development, as well as for the clear, but poorly understood role of LRP1b in cancer.     

Nuclear Receptor CAR Represses TNFα-Induced Cell Death by Interacting with the Anti-Apoptotic GADD45B

Background

Phenobarbital (PB) is the most well-known among numerous non-genotoxic carcinogens that cause the development of hepatocellular carcinoma (HCC). PB activates nuclear xenobiotic receptor Constitutive Active/Androstane Receptor (CAR; NR1I3) and this activation is shown to determine PB promotion of HCC in mice. The molecular mechanism of CAR-mediated tumor promotion, however, remains elusive at the present time. Here we have identified Growth Arrest and DNA Damage-inducible 45β (GADD45B) as a novel CAR target, through which CAR represses cell death.

Methodology/Principal Findings

PB activation of nuclear xenobiotic receptor CAR is found to induce the Gadd45b gene in mouse liver throughout the development of HCC as well as in liver tumors. Given the known function of GADD45B as a factor that represses Mitogen-activated protein Kinase Kinase 7 - c-Jun N-terminal Kinase (MKK7-JNK) pathway-mediated apoptosis, we have now demonstrated that CAR interacts with GADD45B to repress Tumor Necrosis Factor α ( TNFα)-induced JNK1 phosphorylation as well as cell death. Primary hepatocytes, prepared from Car^+/+, Car^−/−, Gadd45b^+/+ and Gadd45b^−/− mice, were treated with TNFα and Actinomycin D to induce phosphorylation of JNK1 and cell death. Co-treatment with the CAR activating ligand TCPOBOP (1,4 bis[2-(3,5-dichloropyridyloxy)]benzene) has resulted in repression of both phosphorylation and cell death in the primary hepatocytes from Car^+/+ but not Car^−/−mice. Repression by TCPOBOP was not observed in those prepared from Gadd45b^−/− mice. In vitro protein-protein interaction and phosphorylation assays have revealed that CAR interacts with MKK7 and represses the MKK7-mediated phosphorylation of JNK1.

Conclusions/Significance

CAR can form a protein complex with GADD45B, through which CAR represses MKK7-mediated phosphorylation of JNK1. In addition to activating the Gadd45b gene, CAR may repress death of mouse primary hepatocytes by forming a GADD45B complex and repressing MKK7-mediated phosphorylation of JNK1. The present finding that CAR can repress cell death via its interaction with GADD45B provides an insight for further investigations into the CAR-regulated molecular mechanism by which PB promotes development of HCC.     

Chorioamnionitis and Patent Ductus Arteriosus: A Systematic Review and Meta-Analysis

Background

Chorioamnionitis has recently been reported as a risk factor for various neonatal diseases, including cerebral palsy, bronchopulmonary dysplasia, and necrotizing enterocolitis, but its effect on patent ductus arteriosus (PDA) is unclear. We performed a systematic review and meta-analysis to evaluate the effect of chorioamnionitis on PDA.

Methods

We searched PubMed, EMBASE, Cochrane Library, and KoreaMed databases using the terms: “intrauterine infection” or “maternal infection” or “antenatal infection” or “chorioamnionitis” or “placenta inflammation” or “placenta pathology” or “neonatal outcome” or “neonatal morbidity” or “PDA or patent ductus arteriosus” or “ductus arteriosus,” and “prematurity” or “very low birth weight infant.” Studies were included if they were randomized controlled trials, case–control studies, or cohort studies that included information relating to chorioamnionitis and PDA.

Results

Among 1,571 studies, a total of 23 studies (17,708 cases) were included in the meta-analysis to analyze the relationship between chorioamnionitis and PDA, except one study that only included PDA requiring surgical ligation. The association between chorioamnionitis and PDA was statistically significant (odds ratio [OR] 1.43; 95% confidence interval [CI] 1.19, 1.72; P < 0.0001). In subgroup analysis, clinical chorioamnionitis was not associated with PDA (OR 1.28; 95% CI 1.00, 1.64, 1.790; P = 0.05), whereas histologic chorioamnionitis (OR 1.54; 95% CI 1.10, 2.15; P = 0.01) and chorioamnionitis diagnosed from both clinical and histologic findings (OR 1.75; 95% CI 1.07, 2.86; P = 0.03) showed significant associations with PDA. Chorioamnionitis did not increase the risk of PDA requiring surgical ligation (OR 1.23; 95% CI 0.69, 2.17; P = 0.48), and antenatal steroid use reduced the risk of PDA (OR 0.62; 95% CI 0.42, 0.90; P = 0.01) after chorioamnionitis.

Conclusions

The results from this meta-analysis support an association between maternal chorioamnionitis and PDA in offspring.     

Thermal niche for in situ seed germination by Mediterranean mountain streams: model prediction and validation for Rhamnus persicifolia seeds

Background and Aims

Mediterranean mountain species face exacting ecological conditions of rainy, cold winters and arid, hot summers, which affect seed germination phenology. In this study, a soil heat sum model was used to predict field emergence of Rhamnus persicifolia, an endemic tree species living at the edge of mountain streams of central eastern Sardinia.

Methods

Seeds were incubated in the light at a range of temperatures (10–25 and 25/10 °C) after different periods (up to 3 months) of cold stratification at 5 °C. Base temperatures (T_b), and thermal times for 50 % germination (θ₅₀) were calculated. Seeds were also buried in the soil in two natural populations (Rio Correboi and Rio Olai), both underneath and outside the tree canopy, and exhumed at regular intervals. Soil temperatures were recorded using data loggers and soil heat sum (°Cd) was calculated on the basis of the estimated T_b and soil temperatures.

Key Results

Cold stratification released physiological dormancy (PD), increasing final germination and widening the range of germination temperatures, indicative of a Type 2 non-deep PD. T_b was reduced from 10·5 °C for non-stratified seeds to 2·7 °C for seeds cold stratified for 3 months. The best thermal time model was obtained by fitting probit germination against log °Cd. θ₅₀ was 2·6 log °Cd for untreated seeds and 2·17–2·19 log °Cd for stratified seeds. When θ₅₀ values were integrated with soil heat sum estimates, field emergence was predicted from March to April and confirmed through field observations.

Conclusions

T_b and θ₅₀ values facilitated model development of the thermal niche for in situ germination of R. persicifolia. These experimental approaches may be applied to model the natural regeneration patterns of other species growing on Mediterranean mountain waterways and of physiologically dormant species, with overwintering cold stratification requirement and spring germination.     

Duplication and diversification of the hypoxia-inducible IGFBP-1 gene in zebrafish

Background

Gene duplication is the primary force of new gene evolution. Deciphering whether a pair of duplicated genes has evolved divergent functions is often challenging. The zebrafish is uniquely positioned to provide insight into the process of functional gene evolution due to its amenability to genetic and experimental manipulation and because it possess a large number of duplicated genes.

Methodology/Principal Findings

We report the identification and characterization of two hypoxia-inducible genes in zebrafish that are co-ortholgs of human IGF binding protein-1 (IGFBP-1). IGFBP-1 is a secreted protein that binds to IGF and modulates IGF actions in somatic growth, development, and aging. Like their human and mouse counterparts, in adult zebrafish igfbp-1a and igfbp-1b are exclusively expressed in the liver. During embryogenesis, the two genes are expressed in overlapping spatial domains but with distinct temporal patterns. While zebrafish IGFBP-1a mRNA was easily detected throughout embryogenesis, IGFBP-1b mRNA was detectable only in advanced stages. Hypoxia induces igfbp-1a expression in early embryogenesis, but induces the igfbp-1b expression later in embryogenesis. Both IGFBP-1a and -b are capable of IGF binding, but IGFBP-1b has much lower affinities for IGF-I and -II because of greater dissociation rates. Overexpression of IGFBP-1a and -1b in zebrafish embryos caused significant decreases in growth and developmental rates. When tested in cultured zebrafish embryonic cells, IGFBP-1a and -1b both inhibited IGF-1-induced cell proliferation but the activity of IGFBP-1b was significantly weaker.

Conclusions/Significance

These results indicate subfunction partitioning of the duplicated IGFBP-1 genes at the levels of gene expression, physiological regulation, protein structure, and biological actions. The duplicated IGFBP-1 may provide additional flexibility in fine-tuning IGF signaling activities under hypoxia and other catabolic conditions.     

Tamoxifen-independent recombination in the RIP-CreER mouse

Background

The inducible Cre-lox system is a valuable tool to study gene function in a spatial and time restricted fashion in mouse models. This strategy relies on the limited background activity of the modified Cre recombinase (CreER) in the absence of its inducer, the competitive estrogen receptor ligand, tamoxifen. The RIP-CreER mouse (Tg (Ins2-cre/Esr1) 1Dam) is among the few available β-cell specific CreER mouse lines and thus it has been often used to manipulate gene expression in the insulin-producing cells of the endocrine pancreas.

Principal Findings

Here, we report the detection of tamoxifen-independent Cre activity as early as 2 months of age in RIP-CreER mice crossed with three distinct reporter strains.

Significance

Evidence of Cre-mediated recombination of floxed alleles even in the absence of tamoxifen administration should warrant cautious use of this mouse for the study of pancreatic β-cells.