Biomedical subjects

Troy L Spilde

Publications and source records attributed to Troy L Spilde.

9 recordsLinked to original sources

PubMed2004

Aberrant fibroblast growth factor receptor 2 signalling in esophageal atresia with tracheoesophageal fistula.

BACKGROUND: Although the pathogenesis of esophageal atresia with tracheoesophageal fistula (EA/TEF) remains unknown, it has been shown that despite its esophageal appearance, the fistula tract originates from respiratory epithelium. The authors now hypothesize that defects in fibroblast growth factor (FGF) signaling contribute to the esophaguslike phenotype of the fistula tract. FGF2R is critical to normal lung morphogenesis and occurs in 2 isoforms (FGF2RIIIb and FGF2RIIIc), each with different ligand-binding specificity. To characterize FGF signaling in the developing EA/TEF, the authors analyzed levels of FGF2R splice variants in experimental EA/TEF. METHODS: The standard Adriamycin-induced EA/TEF model in rats was used. Individual foregut components from Adriamycin-treated and control embryos were processed for real-time, fluorescence-activated semiquantitative reverse transcriptase polymerase chain reaction on gestational days 12.5 and 13.5. RESULTS: Both fistula tract and Adriamycin-treated or normal esophagus showed significantly lower levels of FGF2RIIIb than either Adriamycin-treated lung buds (E12.5, P =.02; E13.5, P <.005) or normal lung buds (E12.5, P <.005; E13.5, P <.01). At E13.5, the fistula tract had lower levels of FGF2RIIIc than either treated (P <.01) or normal lung (P <.05). CONCLUSIONS: Levels of FGF2R in the developing fistula tract resemble that of distal esophagus rather than developing lung. This defect in FGF2RIIIb signaling may account for the nonbranching, esophaguslike phenotype of the fistula, despite its respiratory origin.

Animals↗

PubMed2003

Evaluation of pectus bar position and osseous bone formation.

PURPOSE: Minimally invasive repair has become a popular approach for pectus excavatum (PE). The bar is secured to the thoracic wall and left for approximately 2 years. The authors have noticed an intense bone formation (BF) around some of these bars at removal. A review of children undergoing bar removal was performed to better understand this BF in relation to bar placement. METHODS: A retrospective review of children undergoing bar removal after PE repair since January 1998 was performed. Chart review included age at bar insertion and removal, bar insertion position (subcutaneous [SC] v submuscular [SM]), BF on Chest x-ray and at bar removal, operating time, and estimated blood loss (EBL). RESULTS: Thirty-six patients underwent bar removal during the study period (16 SC and 20 SM). Chest x-ray evaluation was possible in 27 patients (16 SM, 11 SC). No difference existed for length of time the bar was in place or age at insertion/removal between groups. EBL was higher in the SM (18.3 v 8.8 mL, not significant). BF was seen radiographically in 15 SM and 3 SC patients (P <.001). BF was encountered at removal in 19 SM patients and a single SC patient (P <.001). Operating time was statistically longer (P <.01) for the SM group (30.2 v 15.6 min). CONCLUSIONS: Bar position during repair of PE is important. SM positioning virtually always results in BF with increased EBL and statistically longer operating time at removal. Careful placement of the bar in the SC position without violating the fascia should be used to avoid these undesirable effects.

Blood Loss, Surgical↗

PubMed2003

Necrotizing enterocolitis in full-term infants.

OBJECTIVES: Although necrotizing enterocolitis (NEC) is primarily a disease of prematurity, full-term infants account for approximately 10% of cases. Previous studies have reported conflicting results regarding NEC in full-term (FT) versus preterm (PT) infants. A review of all infants diagnosed with NEC at our institution over the past 3 decades was performed to identify factors associated with this disease in full-term neonates. METHODS: The charts of all infants with definitive NEC from January 1, 1972 through January 1, 2001 were reviewed. Two hundred seventy-seven patients made up the study group: 251 PT and 26 FT infants. Data regarding demographics, clinical presentation, management, outcome, and other variables were collected. FT and PT infants were compared. RESULTS: Mean gestational age and birth weight in the FT group were 39.3 weeks and 3,132 g versus 30.2 weeks and 1,396 g for PT infants. Apgar scores were similar. Mean age at diagnosis was 5 days in FT versus 13 days in PT neonates (P <.001). Enteral nutrition was initiated earlier in FT infants (1.6 days v 3.1 days; P <.001), and FT infants were discharged an average of 14 days earlier than PT infants (P value not significant). Factors predisposing to NEC were found in 62% (16 of 26) of patients-heart disease in 6 infants and other conditions in 10 patients. Cardiac disease was found significantly more often (23% v 10%; P =.027) in FT infants. Survival rate was 65% (17 of 26) in the FT group versus 69% (173 of 251) in the PT infants (P value not significant). CONCLUSIONS: FT infants with NEC differ from their PT counterparts in several distinct ways. FT neonates had NEC at a significantly earlier age, perhaps owing to earlier initiation of feeding. There was a correlation between age at which feeding was begun and age of onset of NEC. Additionally, an association between cardiac disease and development of NEC in term infants was shown. Predisposing factors were present in a majority of FT infants. In contrast to other reports, the outcome of NEC in full-term infants was no better than for PT infants.

Age of Onset↗

PubMed2003

Fibroblast growth factor signaling in the developing tracheoesophageal fistula.

BACKGROUND/PURPOSE: The Adriamycin-induced rat model of esophageal atresia and tracheoesophageal fistula (EA/TEF) provides a reliable system for the study of EA/TEF pathogenesis. The authors previously hypothesized that faulty branching lung morphogenesis pathways were a critical component of its pathogenesis. The authors have found evidence for faulty fibroblast growth factor (FGF) signaling related to epithelial-mesenchymal interactions in the fistula tract. To better define FGF signaling, the differential expression of FGF ligands and their receptors between lung, fistula tract, and esophagus are described. METHODS: Time-dated pregnant, Sprague-Dawley rats were injected with Adriamycin (2 mg/kg intraperitoneally) on days 6 through 9 of gestation. Tissues were processed for histology and reverse transcriptase polymerase chain reaction. FGF-1, -7 and -10 were measured from whole lung, fistula tract, and esophagus of TEF or normal embryos. Expression of FGF2RIIIb and FGF2RIIIc receptors was measured in isolated epithelium and mesenchyme of lung and fistula tract of TEF embryos as well as lung and esophagus from normal controls. RESULTS: FGF-1 mRNA was present in the fistula tract and normal and Adriamycin-exposed lung but absent from whole esophagus. Interestingly, FGF-7 mRNA was present only in normal lung. FGF-10 was present in all tissues examined. FGF2RIIIb mRNA was absent in fistula mesenchyme but present in all other tissues examined. However, the splice variant FGF2RIIIc mRNA was present in all tissues examined. CONCLUSIONS: These findings support defective FGF signaling in the rat model of EA/TEF. Absence of FGF-7 mRNA in Adriamycin-exposed tissues suggests the primary effect of Adriamycin may be to inhibit FGF-7 expression. Moreover, absence of FGF2RIIIb in fistula mesenchyme may be caused by loss of positive feedback from FGF-7, its normal obligate ligand. Understanding these specific defects in FGF signaling may provide insight into faulty mechanisms of EA/TEF.

Abnormalities, Drug-Induced↗

PubMed2003

Defective sonic hedgehog signaling in esophageal atresia with tracheoesophageal fistula.

BACKGROUND: The pathogenesis of esophageal atresia and tracheoesophageal fistula (EA/TEF) remains unknown. We have found previously that an initial esophageal atresia, followed by an abnormal (absent) branching pattern of the middle branch of a trifurcation of the lung/tracheal bud, leads to the neonatal finding of TEF. Mice null mutant for hedgehog signaling can experience the development of EA/TEF, but the mechanism for this development is also unknown. Given that EA/TEF in humans appears not to be due to genetic defects, a hedgehog mutation cause seems very unlikely. However, defective hedgehog signaling that is caused by environmental effects in the human embryo likely could be implicated. We studied a teratogen-induced model of EA/TEF to determine the mechanism by which defective hedgehog signaling may lead to EA/TEF. METHODS: We injected Adriamycin into pregnant rats to induce EA/TEF in rat embryos. We first quantified sonic hedgehog (Shh) signaling pathway molecule expression using real-time, semiquantitative reverse-transcriptase polymerase chain reaction for Shh, Shh receptors (patched and smoothened), and downstream intracellular targets of those receptors (Gli family members). On the basis of these findings, we then developed an in vitro culture system for the day-12 embryonic TEF and manipulated Shh signaling using either exogenous Shh or Shh inhibitors. RESULTS: By reverse transcriptase-polymerase chain reaction, a unique difference between the fistula tract and control tissues was that Gli-2 (downstream signaling molecule of Shh) messenger RNA levels were much lower in the fistula tract than in the adjacent esophagus (P =.002). Surprisingly, in the culture experiments, the fistula tract was induced to branch by exogenous Shh. Such branching of the fistula was unexpected and further supports the presumed respiratory origin of the fistula tract because the normal lung, but not normal esophagus, branched in response to Shh. The Shh inhibitor had no effect, which indicated that defective signaling, rather than hyperfunctioning Shh, is critical to the nonbranching phenotype of the fistula tract in TEF. CONCLUSIONS: The recapitulation of respiratory developmental morphogenesis by the fistula tract of TEF in the presence of exogenous Shh, together with the quantitative reduction in normal, endogenous levels of Gli-2, strongly suggests that 1 mechanism for the formation of the fistula tract is the lack of proper Shh signaling because of Gli-2 deficiency, with subsequent straight, nonbranching caudal growth of the fistula tract. This deficiency can be rescued by excess exogenous Shh, thus reestablishing respiratory morphogenesis.

Animals↗

PubMed2002

Complete discontinuity of the distal fistula tract from the developing gut: direct histologic evidence for the mechanism of tracheoesophageal fistula formation.

The embryogenesis of tracheoesophageal anomalies remains controversial. The purpose of this study was to better define the embryogenesis of developing esophageal atresia with tracheoesophageal fistula (EA/TEF), with specific attention to the controversial issue of whether a discontinuity exists in the foregut during its development of EA/TEF. Pregnant outbred rats were injected with adriamycin (2 mg/kg i.p.) on days 6-9 of gestation (E6-E9). At E12.5 and 13.5, microdissection of the entire foregut was performed. Foreguts were examined by phase microscopy, and serial, precisely transverse sections were created for hematoxylin and eosin (H&E) staining. Gross microdissection of the developing foregut at E12.5 (n = 9) revealed a blind-ending, bulbous fistula tract arising from the middle branch of the tracheal trifurcation (as seen by direct and phase microscopy). No connection with the gut could be appreciated at E12.5, but by E13.5 (n = 10) there was an obvious connection between the fistula and the stomach. Serial H&E transverse sections also demonstrated a blind-ending fistula tract arising from the trachea at E12.5. This fistula tract was clearly discontinuous from the developing stomach, which appeared much further caudal to the end of the fistula tract. These results strongly support a model of experimental TEF wherein the fistula tract arises from a trifurcation of the trachea, and (only during a specific gestational window between days 12.5 and 13.5) there is discontinuity between the fistula tract and the stomach. By day 13.5, the fistula joins with the stomach anlage. These observations in the developing EA/TEF should help to resolve the controversy about the mechanism of EA/TEF formation.

Animals↗

PubMed2002

Lectin as a marker for staining and purification of embryonic pancreatic epithelium.

The embryonic pancreatic epithelium, and later the ductal epithelium, is known to give rise to the endocrine and exocrine cells of the developing pancreas, but no specific surface marker for these cells has been identified. Here, we utilized Dolichos Biflorus Agglutinin (DBA) as a specific marker of these epithelial cells in developing mouse pancreas. From the results of an immunofluorescence study using fluorescein-DBA and pancreatic specific cell markers, we found that DBA detects specifically epithelial, but neither differentiating endocrine cells nor acinar cells. We further applied this marker in an immunomagnetic separation system (Dynabead system) to purify these putative multi-potential cells from a mixed developing pancreatic cell population. This procedure could be applied to study differentiation and cell lineage selections in the developing pancreas, and also may be applicable to selecting pancreatic precursor cells for potential cellular engineering.

Animals↗

PubMed2002

Retinoid signaling controls mouse pancreatic exocrine lineage selection through epithelial-mesenchymal interactions.

BACKGROUND & AIMS: The early embryonic pancreas gives rise to exocrine (ducts and acini) and endocrine lineages. Control of exocrine differentiation is poorly understood, but may be a critical avenue through which to manipulate pancreatic ductal carcinoma. Retinoids have been shown to change the character of pancreatic ductal cancer cells to a less malignant phenotype. We have shown that 9-cis retinoic acid (9cRA) inhibits acinar differentiation in the developing pancreas, in favor of ducts, and we wanted to determine the role of retinoids in duct versus acinar differentiation. METHODS: We used multiple culture systems for the 11-day embryonic mouse pancreas. RESULTS: Retinoic acid receptor (RAR)-selective agonists mimicked the acinar suppressive effect of 9cRA, suggesting that RAR-RXR heterodimers were critical to ductal differentiation. RARalpha was only expressed in mesenchyme, whereas RXRalpha was expressed in epithelium and mesenchyme. Retinaldehyde dehydrogenase 2, a critical enzyme in retinoid synthesis, was expressed only in pancreatic epithelium. 9cRA did not induce ductal differentiation in the absence of mesenchyme, implicating a requirement for mesenchyme in 9cRA effects. Mesenchymal laminin is necessary for duct differentiation, and retinoids are known to enhance laminin expression. In 9cRA-treated pancreas, immunohistochemistry for laminin showed a strong band of staining around ducts, and blockage of laminin signaling blocked all 9cRA effects. Western blot and RT-PCR of pancreatic mesenchyme showed laminin-beta1 protein and mRNA induction by 9cRA. CONCLUSIONS: Retinoids regulate exocrine lineage selection through epithelial-mesenchymal interactions, mediated through up-regulation of mesenchymal laminin-1.

Alitretinoin↗

PubMed2002

Thyroid transcription factor-1 expression in the human neonatal tracheoesophageal fistula.

BACKGROUND/PURPOSE: Esophageal atresia with tracheoesophageal fistula (EA/TEF) is a relatively common congenital anomaly, but its pathogenesis remains poorly understood. Previous studies using the experimental Adriamycin-induced rat model of EA/TEF suggest that the fistula tract, or "distal esophagus," is derived from respiratory epithelium and expresses the respiratory-specific transcription factor TTF-1. To better correlate the experimental rat model of EA/TEF with the human anomaly, we looked for evidence of a respiratory-derived origin in the neonatal TEF. METHODS: After IRB approval, 2 human TEF samples were removed at the time of surgery. The tissue from the fistula tract was trimmed in accordance with what the surgeons deemed to be appropriate for the preparation for a primary anastomosis. The tissues then were processed for reverse transcription polymerase chain reaction (RT-PCR), histology, and immunohistochemistry. Normal embryonic lung cDNA was used for positive controls. RESULTS: Histologic examination of tissue specimens showed many epithelial tubules within loose connective tissue and a disorganized muscular coat. Thyroid transcription factor one (TTF-1) and hepatocyte nuclear factor 3beta (HNF-3beta) were shown to be present in the tissue specimen by RT-PCR and immunohistochemistry. In addition, FGF-10, a strong morphogen present in the developing lung, and FGF-1 were both present by RT-PCR. The PCR band sizes for both FGF-1 and -10 were appropriate, using human embryonic cDNA as a control, and the bands were confirmed as nongenomic by either placing the PCR primers across a known intron sequence (TTF-1) or by absence of a band in a negative RT control (HNF-3beta, FGF-10, FGF-1). CONCLUSIONS: The presence of TTF-1 in the neonatal TEF shows, for the first time, that parallels may be drawn between the experimental rat model of TEF and the human anomaly at the molecular level. Moreover, these results suggest that the fistula tract in the human neonate is derived from a respiratory cell lineage. This respiratory origin of the human TEF may explain the poor esophageal motility (and subsequent serious respiratory complications) of the distal segment after standard EA/TEF repair.

Humans↗