Development Of Exocrine Pancreas Research Articles

Zebrafish reporter lines reveal in vivo signaling pathway activities involved in pancreatic cancer

Pancreatic adenocarcinoma, one of the worst malignancies of the exocrine pancreas, is a solid tumor with increasing incidence and mortality in industrialized countries. This condition is usually driven by oncogenic KRAS point mutations and evolves into a highly aggressive metastatic carcinoma due to secondary gene mutations and unbalanced expression of genes involved in the specific signaling pathways. To examine in vivo the effects of KRASG12D during pancreatic cancer progression and time correlation with cancer signaling pathway activities, we have generated a zebrafish model of pancreatic adenocarcinoma in which eGFP-KRASG12D expression was specifically driven to the pancreatic tissue by using the GAL4/UAS conditional expression system. Outcrossing the inducible oncogenic KRASG12D line with transgenic zebrafish reporters, harboring specific signaling responsive elements of transcriptional effectors, we were able to follow TGFβ, Notch, Bmp and Shh activities during tumor development. Zebrafish transgenic lines expressing eGFP-KRASG12D showed normal exocrine pancreas development until 3 weeks post fertilization (wpf). From 4 to 24 wpf we observed several degrees of acinar lesions, characterized by an increase in mesenchymal cells and mixed acinar/ductal features, followed by progressive bowel and liver infiltrations and, finally, highly aggressive carcinoma. Moreover, live imaging analysis of the exocrine pancreatic tissue revealed an increasing number of KRAS-positive cells and progressive activation of TGFβ and Notch pathways. Increase in TGFβ, following KRASG12D activation, was confirmed in a concomitant model of medulloblastoma (MDB). Notch and Shh signaling activities during tumor onset were different between MDB and pancreatic adenocarcinoma, indicating a tissue-specific regulation of cell signaling pathways. Moreover, our results show that a living model of pancreatic adenocarcinoma joined with cell signaling reporters is a suitable tool for describing in vivo the signaling cascades and molecular mechanisms involved in tumor development and a potential platform to screen for novel oncostatic drugs.

Multiple Ribosomal Proteins Are Expressed at High Levels in Developing Zebrafish Endoderm and Are Required for Normal Exocrine Pancreas Development

Ribosomal protein L (rpl) genes are essential for assembly of the 60S subunit of the eukaryotic ribosome and may also carry out additional extra-ribosomal functions. We have identified a common expression pattern for rpl genes in developing zebrafish larvae. After initially widespread expression in early embryos, the expression of multiple rpl genes becomes increasingly restricted to the endoderm. With respect to the pancreas, rpl genes are highly expressed in ptf1a-expressing pancreatic progenitors at 48 hpf, suggesting possible functional roles in pancreatic morphogenesis and/or differentiation. Utilizing two available mutant lines, rpl23a(hi2582) and rpl6(hi3655b), we found that ptf1a-expressing pancreatic progenitors fail to properly expand in embryos homozygous for either of these genes. In addition to these durable homozygous phenotypes, we also demonstrated recoverable delays in ptf1a-expressing pancreatic progenitor expansion in rpl23a(hi2582) and rpl6(hi3655b) heterozygotes. Disruptions in ribosome assembly are generally understood to initiate a p53-dependent cellular stress response. However, concomitant p53 knockdown was unable to rescue normal pancreatic progenitor expansion in either rpl23a(hi2582) or rpl6(hi3655b) mutant embryos, suggesting required and p53-independent roles for rpl23a and rpl6 in pancreas development.

Def Functions as a Cell Autonomous Factor in Organogenesis of Digestive Organs in Zebrafish

Digestive organs originate from the endoderm. Morphogenesis of the digestive system is precisely controlled by multiple factors that dictate the cell fate and behavior so that the specific digestive organs are timely formed in the right place and develop into right size and structure. We showed previously that digestive organ expansion factor (def) is a gene whose expression is enriched in the liver, pancreas and intestine. Loss-of-function of def in the defhi429 mutant confers hypoplastic digestive organs partly due to alteration of expression of genes related to the p53 pathway. However, the molecular mechanism for the involvement of Def in the organogenesis of digestive organs is still largely unknown. For example, it is not known whether Def regulates specific pathways in a specific organ. To address this question, we generated four independent Tg(fabp10a:def) transgenic fish lines which over-expressed Def specifically in the liver. We characterized Tg-I, one of the transgenic lines, in detail with genetic, molecular and histological approaches. We found that Tg-I restored the liver but not exocrine pancreas and intestine development in the defhi429 mutant. However, Tg-I adult fish in the wild type (WT) background exhibits reduced liver-to-body ratio and all four transgenic lines conferred abnormal intrahepatic structure. Microarray data analysis showed that certain specific functional pathways were affected in the liver of Tg-I. These results demonstrate that Def functions in a cell autonomous manner during early liver development and aberrant Def protein expression might lead to disruption of the structural integrity of a normal adult liver.

Rbms3, an RNA-Binding Protein, Mediates the Expression ofPtf1aby Binding to Its 3′UTR During Mouse Pancreas Development

The development of the pancreas is a complicated process that is regulated on several levels. Pancreas transcription factor 1, alpha subunit (Ptf1a), also known as p48, is a pancreas-specific basic helix-loop-helix transcription factor that is critical for both exocrine pancreas development and maintenance of acinar cell differentiation. Based on a differential screening assay, we identified Rbms3, a gene encoding a glycine-rich RNA-binding protein, to be specifically expressed in the neural tube and the pancreatic rudiment of e10.5 embryos. The presence of Rbms3 in the early developing pancreas suggests that specific post-transcriptional regulation mechanisms play an important role in controlling pancreas development. In this study, we show that Rbms3 binds to the 3'UTR of Ptf1a mRNA, but not the 3'UTR of Pdx1, which is another pancreatic transcription factor. The ectopic expression of Rbms3 stimulates the translation of a reporter gene carrying the Ptf1a 3'UTR. In addition, when Rbms3 expression is suppressed in the AR42J-B13 pancreatic exocrine cell line, the expression of Ptf1a is also down-regulated. These results suggest that binding of Rbms3 to the 3'UTR of Ptf1a regulates the production of the Ptf1a protein and, thereby, indirectly regulates the expression of the Ptf1a downstream target genes.

TRPM7 and TRPM8 Ion Channels in Pancreatic Adenocarcinoma: Potential Roles as Cancer Biomarkers and Targets

Transient receptor potential (TRP) ion channels are essential for normal functions and health by acting as molecular sensors and transducing various stimuli into cellular and physiological responses. Growing evidence has revealed that TRP ion channels play important roles in a wide range of human diseases, including malignancies. In light of recent discoveries, it has been found that TRP melastatin-subfamily members, TRPM7 and TRPM8, are required for normal and cancerous development of exocrine pancreas. We are currently investigating the mechanisms which mediate the functional roles of TRPM7 and TRPM8 and attempting to develop these ion channels as clinical biomarkers and therapeutic targets for achieving the goal of personalized therapy in pancreatic cancer.

Histone deacetylase 1 is required for exocrine pancreatic epithelial proliferation in development and cancer

Histone deacetylases (HDACs) play important roles in the epigenetic control of development, and aberrant expression of HDACs has been implicated in human diseases including cancer. Among the mammalian HDACs, HDAC1 has been extensively studied, but its role in exocrine pancreatic morphogenesis and cancer is still poorly understood. The goal of this study is to determine the functional role of HDAC1 in normal development of exocrine pancreas using zebrafish as the model organism as well as in human pancreatic adenocarcinoma. The zebrafish germline loss-of-function mutation hdac1hi1618 caused impaired cell cycle progression in pancreatic epithelia, resulting in growth arrest and dysmorphogenesis of exocrine pancreas. In human pancreatic adenocarcinoma tissues and cell lines, HDAC1 was expressed at variably elevated levels. RNA interference-induced silencing of HDAC1 diminished proliferation of the cancer cells and cell cycle progression. The proliferative arrest in the developing exocrine pancreas and pancreatic cancer cells was associated with up-regulated expression of the cyclin-dependent kinase inhibitors and the sonic hedgehog signaling components. This study indicates that HDAC1 is required for pancreatic epithelial proliferation in development and cancer. We hypothesize that aberrant expression of HDAC1 modulates the developmental and signaling pathways in exocrine pancreatic epithelia and consequently the genes required for cellular proliferation during development and progression of pancreatic neoplasia.

Genetic inducible fate mapping in larval zebrafish reveals origins of adult insulin-producing β-cells

The Notch-signaling pathway is known to be fundamental in controlling pancreas differentiation. We now report on using Cre-based fate mapping to indelibly label pancreatic Notch-responsive cells (PNCs) at larval stages and follow their fate in the adult pancreas. We show that the PNCs represent a population of progenitors that can differentiate to multiple lineages, including adult ductal cells, centroacinar cells (CACs) and endocrine cells. These endocrine cells include the insulin-producing β-cells. CACs are a functional component of the exocrine pancreas; however, our fate-mapping results indicate that CACs are more closely related to endocrine cells by lineage as they share a common progenitor. The majority of the exocrine pancreas consists of the secretory acinar cells; however, we only detect a very limited contribution of PNCs to acinar cells. To explain this observation we re-examined early events in pancreas formation. The pancreatic anlage that gives rise to the exocrine pancreas is located in the ventral gut endoderm (called the ventral bud). Ptf1a is a gene required for exocrine pancreas development and is first expressed as the ventral bud forms. We used transgenic marker lines to observe both the domain of cells expressing ptf1a and cells responding to Notch signaling. We do not detect any overlap in expression and demonstrate that the ventral bud consists of two cell populations: a ptf1-expressing domain and a Notch-responsive progenitor core. As pancreas organogenesis continues, the ventral bud derived PNCs align along the duct, remain multipotent and later in development differentiate to form secondary islets, ducts and CACs.

Exocrine Pancreas Development and Trypsin Expression in Cultured European Sea Bass (Dicentrarchus labrax) Larvae

The ontogenesis and formation stages of exocrine pancreas in European sea bass (D. labrax) larvae were investigated from hatching to 40 days after hatching (DAH). Histological and enzymatical techniques were used to explain the functional development of the pancreas in D. labrax with the expression of trypsinogen activity. The incipient pancreas appeared as a lamination of the dorsal wall of the digestive tract. It was observed that the primary visible indication of exocrine cell differentiation was polarization. The first zymogen granules and pancreas with exocrine polyhedral cells appeared on 6 DAH and became abundant as a compact structure located dorsal and slightly posterior to the liver. At the same time, firstly, anus and then mouth were opened, and total lengths of larvae were determined as 3.47±0.26 mm. Until larval metamorphosis, the pancreas became diffuse, spreading throughout the mesentery enclosure, the stomach, the upper intestine and the pyloric caeca. On the other hand, zymogen granules were more numerous and larger, and a greater quantity of material was carried by the ducts, indicating an increased cellular activity. The specific activity of trypsin was determined as early as after hatching (42.54±6.8 mU/mg protein-1) at 4.28±0.2 mm total length of larvae and increased immediately during the following days especially after exogenous feeding. The highest tryptic activity was detected on 30 DAH as 122.45±11.76 mU/mg protein-1. It is concluded that exocrine pancreas organogenesis is the main critical step of the zymogen granules and trypsin activity is present as early as after hatching and continuously increasing with larval period of D. labrax.

Exdpf Is a Key Regulator of Exocrine Pancreas Development Controlled by Retinoic Acid and ptf1a in Zebrafish

Both endocrine and exocrine pancreatic cells arise from pancreatic-duodenal homeobox 1 (pdx1)-positive progenitors. The molecular mechanisms controlling cell fate determination and subsequent proliferation, however, are poorly understood. Unlike endocrine cells, less is known about exocrine cell specification. We report here the identification and characterization of a novel exocrine cell determinant gene, exocrine differentiation and proliferation factor (exdpf), which is highly expressed in the exocrine cell progenitors and differentiated cells of the developing pancreas in zebrafish. Knockdown of exdpf by antisense morpholino caused loss or significant reduction of exocrine cells due to lineage-specific cell cycle arrest but not apoptosis, whereas the endocrine cell mass appeared normal. Real-time PCR results demonstrated that the cell cycle arrest is mediated by up-regulation of cell cycle inhibitor genes p21Cip, p27Kip, and cyclin G1 in the exdpf morphants. Conversely, overexpression of exdpf resulted in an overgrowth of the exocrine pancreas and a severe reduction of the endocrine cell mass, suggesting an inhibitory role for exdpf in endocrine cell progenitors. We show that exdpf is a direct target gene of pancreas-specific transcription factor 1a (Ptf1a), a transcription factor critical for exocrine formation. Three consensus Ptf1a binding sites have been identified in the exdpf promoter region. Luciferase assay demonstrated that Ptf1a promotes transcription of the exdpf promoter. Furthermore, exdpf expression in the exocrine pancreas was lost in ptf1a morphants, and overexpression of exdpf successfully rescued exocrine formation in ptf1a-deficient embryos. Genetic evidence places expdf downstream of retinoic acid (RA), an instructive signal for pancreas development. Knocking down exdpf by morpholino abolished ectopic carboxypeptidase A (cpa) expression induced by RA. On the other hand, exdpf mRNA injection rescued endogenous cpa expression in embryos treated with diethylaminobenzaldehyde, an inhibitor of RA signaling. Moreover, exogenous RA treatment induced anterior ectopic expression of exdpf and trypsin in a similar pattern. Our study provides a new understanding of the molecular mechanisms controlling exocrine cell specification and proliferation by a novel gene, exdpf. Highly conserved in mammals, the expression level of exdpf appears elevated in several human tumors, suggesting a possible role in tumor pathogenesis.

Tumour suppressor menin is essential for development of the pancreatic endocrine cells

Mutations of the multiple endocrine neoplasia type 1 (MEN1) gene predispose patients to MEN1 that affects mainly endocrine tissues, suggesting important physiological functions of the gene in adult endocrine cells. Homozygous disruption of Men1 in mice causes embryonic lethality, whereas the eventual involvement of the gene in embryonic development of the endocrine cells remains unknown. Here, we show that homozygous Men1 knockout mice demonstrate a reduced number of glucagon-positive cells in the E12.5 pancreatic bud associated with apoptosis, whereas the exocrine pancreas development in these mice is not affected. Our data suggest that menin is involved in the survival of the early pancreatic endocrine cells during the first developmental transition. Furthermore, chimerism assay revealed that menin has an autonomous and specific effect on the development of islet cells. In addition, using pancreatic bud culture mimicking the differentiation of alpha- and beta-cells during the second transition, we show that loss of menin leads to the failure of endocrine cell development, altered pancreatic structure and a markedly decreased number of cells expressing neurogenin 3, indicating that menin is also required at this stage of the endocrine pancreas development. Taken together, our results suggest that menin plays an indispensable role in the development of the pancreatic endocrine cells.

Conditional ablation of Notch signaling in pancreatic development

The role of the Notch signaling members Notch1, Notch2 and Rbpj in exocrine pancreatic development is not well defined. We therefore analyzed conditional pancreas-specific Rbpj and combined Notch1/Notch2 knockout mice using Ptf1a(+/Cre(ex1)) mice crossed with floxed Rbpj or Notch1/Notch2 mice. Mice were analyzed at different embryonic stages for pancreatic exocrine and endocrine development. The absence of Rbpj in pancreatic progenitor cells impaired exocrine pancreas development up to embryonic day 18.5 and led to premature differentiation of pancreatic progenitors into endocrine cells. In Rbpj-deficient pancreata, amylase-expressing acini and islets formed during late embryonic and postnatal development, suggesting an essential role of Rbpj in early but not late development. Contrary to this severe phenotype, the concomitant inactivation of Notch1 and Notch2 only moderately disturbed the proliferation of pancreatic epithelial cells during early embryonic development, and did not inhibit pancreatic development. Our results show that, in contrast to Rbpj, Notch1 and Notch2 are not essential for pancreatogenesis. These data favor a Notch-independent role of Rbpj in the development of the exocrine pancreas. Furthermore, our findings suggest that in late stages of pancreatic development exocrine cell differentiation and maintenance are independent of Rbpj.

Cloning, expression, and functional characterization of zebrafish Mist1

The basic helix–loop–helix (bHLH) protein Mist1 is an important exocrine pancreas transcriptional factor expressed in the acinar cells of mammals. In the present study, we cloned the homologous Mist1 cDNA encoding a predicted protein of 184 amino acids in zebrafish. The typical bHLH domain of zebrafish Mist1 shares high identity with that of its orthologs in mouse, rat, and human. Expression analysis revealed that Mist1 maternal transcripts are distinct in the very beginning of embryogenesis and that endogenous Mist1 is chronologically expressed in polster, hatching gland, hindbrain and appears exclusively in the pancreas from 72 hpf onward. Knockdown of Mist1 conditionally causes mild morphological defects in embryos. In MO-treated embryos, midbrain–hindbrain boundary is missing and exocrine pancreas is significantly reduced and disorganized. These results suggest that Mist1 functions in an evolutionary conserved way as a key transcriptional regulator specific for exocrine pancreas development in zebrafish.

The bHLH Domain of Mist1 Is Sufficient to Activate Gene Transcription

Mist1 is a tissue-specific basic helix-loop-helix (bHLH) transcription factor that plays an essential role in maintaining and organizing the exocrine pancreas. Consequently, mice lacking Mist1 exhibit disrupted acinar cellular polarity and defective zymogen granule trafficking. Despite extensive studies demonstrating a requirement for Mist1 in exocrine pancreas development and function, little is known about the molecular targets for Mist1 interaction and the mechanism(s) of how Mist1 regulates gene transcription. To address these deficiencies, a series of molecular studies was performed to identify the preferred Mist1 dimer complex and to establish the preferred DNA binding site for this bHLH factor. In vivo coimmunoprecipitation assays confirmed that the functional Mist1 complex in pancreatic acinar cells was a Mist1 homodimer that bound to a unique DNA target site known as the TA-E-box. Binding of Mist1 to a TA-E-box-regulated promoter led to transcriptional activation of the target gene. Surprisingly, Mist1 truncations containing only the central bHLH domain retained approximately 80% of transcriptional activity. Coimmunoprecipitation studies demonstrated that the bHLH domain interacted with coactivators belonging to the p300/CBP family, suggesting that Mist1 activates exocrine-specific gene transcription through an acetylation mechanism.

Analyses of pancreas development by generation of gfp transgenic zebrafish using an exocrine pancreas-specific elastaseA gene promoter

In contrast to what we know on development of endocrine pancreas, the formation of exocrine pancreas remains poorly understood. To create an animal model that allows observation of exocrine cell differentiation, proliferation, and morphogenesis in living animals, we used the zebrafish elastaseA ( elaA) regulatory sequence to develop transgenic zebrafish that display highly specific exocrine pancreas expression of GFP in both larvae and adult. By following GFP expression, we found that the pancreas in early development was a relatively compact organ and later extended posterior along the intestine. By transferring the elaA:gfp transgene into slow muscle omitted mutant that is deficient in receiving Hedgehog signals, we further showed that Hedgehog signaling is required for exocrine morphogenesis but not for cell differentiation. We also applied the morpholino knockdown and toxin-mediated cell ablation approaches to this transgenic line. We showed that the development of exocrine pancreas is Islet-1 dependent. Injection of the diphtheria toxin A ( DTA) construct under the elastaseA promoter resulted in selective ablation of exocrine cells while the endocrine cells and other endodermal derivatives (liver and intestine) were not affected. Thus, our works demonstrated the new transgenic line provided a useful experimental tool in analyzing exocrine pancreas development.

Exocrine pancreas development in zebrafish

Although many of the genes that regulate development of the endocrine pancreas have been identified, comparatively little is known about how the exocrine pancreas forms. Previous studies have shown that exocrine pancreas development may be modeled in zebrafish. However, the timing and mechanism of acinar and ductal differentiation and morphogenesis have not been described. Here, we characterize zebrafish exocrine pancreas development in wild type and mutant larvae using histological, immunohistochemical and ultrastructural analyses. These data allow us to identify two stages of zebrafish exocrine development. During the first stage, the exocrine anlage forms from rostral endodermal cells. During the second stage, protodifferentiated progenitor cells undergo terminal differentiation followed by acinar gland and duct morphogenesis. Immunohistochemical analyses support a model in which the intrapancreatic ductal system develops from progenitors that join to form a contiguous network rather than by branching morphogenesis of the pancreatic epithelium, as described for mammals. Contemporaneous appearance of acinar glands and ducts in developing larvae and their disruption in pancreatic mutants suggest that common molecular pathways may regulate gland and duct morphogenesis and differentiation of their constituent cells. By contrast, analyses of mind bomb mutants and jagged morpholino-injected larvae suggest that Notch signaling principally regulates ductal differentiation of bipotential exocrine progenitors.

Pluripotent Unrestricted Somatic Stem Cells (USSC) from Cord Blood Generate Hepatocytes in Vivo, but In Vitro Convert into Immature Endodermal Precursors: The Critical Role of the Biological Niche.

As shown recently by our group (Koegler et al. 2004; J. Exp. Med. 200: 19:1–13) transplantation of USSC in a non-injury model, the preimmune fetal sheep, resulted in more than 20% albumin-producing human parenchymal hepatic cells with absence of cell fusion. Here we examined whether we are able to trigger USSC in vitro into the an endodermal differentiation pathway applying protocols described for both embryonal as well as adult stem cell. The following matrix/growth factors/organic substances were applied: Neurobasal medium/supplement B27/bFGF with and without nicotinamid, on fibronectin, laminin or matrigel for differentiation into the pancreatic development as well as HGF, NGF, bFGF and HGF, FGF4 with or without Oncostatin M to induce liver development. 15 different USSC lines were differentiated for 1, 2, 3, 4 and 6 weeks. Primers were designed with the strategy to define stages of endodermal development on the basis of the embryonic cell development from mouse and human. Therefore the following markers were established: GATA 4, HNF-1, HNF3ß and HNF 4a to define the embryonic and visceral (extra-embryonic) endoderm, a common precursor phenotype for both liver/exocrine pancreas development. To further assess differentiation into liver cells, a-1 antitrypsin, a-fetoprotein, albumin, HGF, Cyp2B6, Cyp3A4, Gys2 and PDX-1, PAX4, ISL-1, NKx6.1, NeuroD, insulin to determine differentiation into the pancreatic development as well as epithelial markers as CK8, CK18, CK19 were analyzed by RT-PCR and subsequent hybridization using gene-specific oligonucleotide probes. USSC were tested negativ for the majority of the markers, only HGF and cytokeratin markers CK8/18 and CK19 were tested positiv. In vitro differentiation shows that USSC never expressed Neuro D, HNF1, HNF3b, PDX-1, PAX4, insulin and a-fetoprotein, but do express depending on the culture conditions common endodermal precursor markers HNF4a, GATA4 (but not HNF1 or HNF3ß) as well as albumin, Cyp2B6, Cyp3A4, Gys2 (liver development) and Nkx6.1 and ISL-1 (pancreatic development. It is very interesting to note that ISL-1, required for the formation of the dorsal mesenchym and essential for the dorsal exocrine pancreas development is strong expressed, although all other markers as PDX-1 and PAX4 were always tested negative. We have never observed the expression of a-fetoprotein, which might be explained by the kinetic of this factor (Young-Yang, Nat.Cell Biology 2004). In summary, the results show that under the vitro conditions applied, only an endodermal precursor phenotype could be established. Therefore at present co-cultivation experiments with injured hepatocytes from sheep/rat/human are performed to mimic the biological niche in vitro as shown by Young-Yang et al. Western blots with an human albumin specific antibody (moAb, clone HAS-11, Sigma) revealed that this approach resulted in an strong expression of albumin in USSC. Moreover, this co-cultivation model is useful for the identification and characterization of factors who are responsible for the biological niche in vivo.

Trypsinogen expression during the development of the exocrine pancreas in winter flounder (Pleuronectes americanus)

Trypsinogen expression during the development of the exocrine pancreas in winter flounder (Pleuronectes americanus)

Differential requirement for ptf1a in endocrine and exocrine lineages of developing zebrafish pancreas

Mammalian studies have implicated important roles for the basic helix–loop–helix transcription factor Ptf1a-p48 in the development of both exocrine and endocrine pancreas. We have cloned the Ptf1a-p48 ortholog in Danio rerio. Early zebrafish ptf1a expression is observed in developing hindbrain and in endodermal pancreatic precursors. Analysis of ptf1a and insulin expression reveals a population of exocrine precursors that, throughout early development, are temporally and spatially segregated from endocrine elements. Morpholino-mediated knockdown of ptf1a confirms early divergence of these endocrine and exocrine lineages. Ptf1a morphants lack differentiated exocrine pancreas, but maintain normal differentiation and organization of the principal islet. In addition to the exocrine phenotype, ptf1a knockdown also reduces the prevalence of a small population of anterior endocrine cells normally found outside the principal islet. Together, these findings suggest the presence of distinct ptf1a-dependent and ptf1a-independent precursor populations in developing zebrafish pancreas.

Digestive enzyme activity during larval development of the Senegal sole (Solea senegalensis)

The activities of the main digestive enzymes (proteases, amylase, lipase) as well as those of acid and alkaline phosphatases were assessed during the larval development of the Senegal sole, Solea senegalensis. Important variations in specific activities of all the enzymes were observed during the period of study and were mostly related to the beginning or the end of metamorphosis. Both acid and alkaline protease activities were identified at early stages of development. Acid proteases accounted for only 10% of total protease activity in a 9 days-old larva, but exceeded 75% in a 33 days-old individual. Maximum lipase activity was related to the development of exocrine pancreas (days 6 to 10) and to metamorphosis. It is suggested this can be related to the metabolism of lipid reserves taking place during this morphological change. Activity of alkaline phosphatase decreased from day 5 to day 20 but therafter revealed a sharp increase, possibly linked to the development of enterocytes. The pattern of development of the main digestive enzymes found in S. senegalensis is similar to that described in other flatfish species.

Postnatal Development of Liver and Exocrine Pancreas in Polycythemic Newborn Infants

SummaryIn 35 newborn infants appropriate for gestational age the influence of neonatal polycythemia (venous hematocrit >60% measured between the second and fourth hour of life) on development of enterohepatic circulation of bile acids, activities of pancreatic enzymes in duodenal juice, and the effects of hemodilution were studied during the second week of life. A significant correlation was found between the initial hematocrit and both the bile acid concentration in serum and lipase and trypsin activity in duodenal juice. Of 35 infants, 10 were not treated with hemodilution due to asymptomatic polycythemia; they had the highest concentration of serum bile acids associated with the lowest lipase and trypsin activity in duodenal juice. However, the 25 infants treated with hemodilution also showed serum bile acid concentrations and lipase and trypsin activity in duodenal juice out of the normal range when compared to normocythemic infants. These data indicate that, during the first days of life, polycythemia results in a delayed postnatal development of enterohepatic circulation of bile acids and exocrine pancreas functions independently from the occurrence of clinical symptoms. Thus, it can be concluded that, on the first day of life, all polycythemic infants should be treated with hemodilution. Moreover, the nutritional management of these infants must also account for the limited functional capacity of the gastrointestinal tract.