Unsuspected transection of lung by pigtail catheter in a premature infant.

Thoracostomy tubes are commonly required to treat pnuemothoraces in premature infants. Evidence of impalement of the lungs by tube thoracostomy has been seen in autopsy studies. In neonates, there has been described a surprisingly high incidence of lung perforation. The premature lung is thought to be at greater risk for this complication owing to the pliant, thin chest wall, the proximity of vital tissues and the fragility of the lung tissue itself. The modified Fuhrman catheter, or polyurethane pigtail catheter, has been developed for the drainage of pneumothorax in premature infants. In a study of complications of the placement of pigtail catheters, no instance of penetration of the lungs was reported. We report the case of a premature infant with pigtail catheter placement that, at autopsy, was found to have impaled the lung and discuss the incidence of lung injury associated with invasive management of pnuemothoraces.

Morphologic changes in alveolar macrophages in response to UVEC-activated pulmonary Type II epithelial cells.

We hypothesize that Type II epithelial cells, which line the distal airspaces of the lung, are early responders to invading pathogens and release a signal, which activates and alters the phenotype and phagocytosis properties of alveolar macrophages even at a distance. The T(7) cell line is a conditionally immortalized murine Type II epithelial cell line developed in our laboratory. Using an in vitro transwell model we have previously shown that UV-irradiated Escherichia coli (UVEC)-stimulated T(7) cells cultured in the lower transwell chamber, release a diffusible signal which activates MH-S cells (immortalized murine alveolar macrophages) cultured in the upper transwell chamber, to produce nitric oxide. Using scanning electron microscopy, we show that MH-S cells activated in this manner exhibit increased cell surface ruffling, numerous long filopodia, increased lamellipodia and cell flattening. DynaBead uptake studies show that these morphologic changes are accompanied by increased phagocytosis. These findings indicate that a diffusible signal released at a distance by UVEC-stimulated Type II epithelial cells initiates changes in morphology and phagocytosis reflective of macrophage activation concomitant with the functional activation we previously reported.

Diffusible signal to murine alveolar macrophages from lipopolysaccharide- and Escherichia coli-stimulated lung Type II epithelial cells.

OBJECTIVE AND DESIGN: To demonstrate a diffusible intercellular macrophage activation factor secreted by Type II alveolar epithelial cells (AECs) in transwell co-cultures. MATERIALS: T(7), our Type II conditionally immortalized AEC line; MH-S, an alveolar macrophage cell line; Lipopolysaccharide (LPS) or uv-killed Escherichia coli (UVEC) for antigen presentation. METHODS: LPS or UVEC stimulation of T(7) cells in the lower chamber was investigated for ability to activate MH-S cells in the upper chamber, as assayed by nitric oxide production and western blots for inducible nitric oxide synthase-2. RESULTS: Both transwell and UVEC-conditioned medium experiments showed secretion of an MH-S activation factor by T(7) cells. Many common inflammatory cytokines were ruled out as this immunoactivator. CONCLUSION: Demonstration of a diffusible activation factor produced by Type II AECs supports their potential role as first responders of innate immunity in the lung.

Atelectasis--an unusual and late complication of lung transplant.

We report a previously unrecognized late complication of allograft lung transplantation - persistent recurrent atelectasis of the transplanted lung. The patient developed sudden, severe respiratory distress about 2 yr after a right lung transplant, because of acute atelectasis of her transplanted lung. Multiple transbronchial biopsies at the time revealed minimal inflammation and no evidence of rejection. She was treated with surfactant replacement therapy, and her collapsed lung fully expanded following surfactant installation. To eliminate the possibility of acquired deficiency of surfactant lipids or proteins, ultrastructural examination and immunostains for surfactant proteins were performed in a transbronchial lung biopsy. No deficiency of surfactant lipids or proteins was found. On ultrastructural examination of the lung biopsy, the number of Type II cells per alveolus and the number of lamellar bodies per square micron of Type II cell cross-sectional area was increased compared with an age-matched control. We conclude that synthesis of surfactant lipids and proteins was unimpaired and because of the patient's response to surfactant replacement therapy, that the increase in number of lamellar bodies could reflect a compensatory mechanism for a surfactant functional defect. The patient later developed breast carcinoma to which she succumbed. We raise the possibility that the functional surfactant defect is a hitherto unrecognized non-metastatic manifestation of malignancy.

Differential expression of VEGF isoforms in mouse during development and in the adult.

Vascular endothelial growth factor (VEGF), a factor that is critical for development of the vascular system in mouse embryos, exists as at least three isoforms, VEGF120, VEGF164, and VEGF188. The isoforms have different affinities for heparan sulfate as well as for the three known VEGF receptors, VEGFR-1 (Flt-1), VEGFR-2 (Flk-1), and neuropilin-1, suggesting that different VEGF isoforms may play distinct roles in vascular development. To determine whether there are differences in the organ-specific expression patterns that would support this concept, we used a quantitative RNase protection assay (RPA) to determine the distribution of different VEGF isoform mRNA in developing and adult mouse organs. Results revealed that the ratios of the three VEGF isoforms changed during organ development and that adult organs expressed different levels of the three VEGF isoforms. Because the lung expressed the highest levels of VEGF188 isoform, we used VEGF isoform-specific in situ hybridization in the developing lung and determined that type II alveolar epithelial cells were expressing high levels of VEGF188 mRNA. Finally, targeted exon deletion of the VEGF gene revealed that mice that developed in the absence of the heparan sulfate binding isoforms VEGF164 and VEGF188, displayed a variety of vascular defects, including abnormal pulmonary vascular development. Our results support the concept that different VEGF isoforms have distinct functions in vascular development.

Generation of an immortal differentiated lung type-II epithelial cell line from the adult H-2K(b)tsA58 transgenic mouse.

This paper describes a new fully differentiated Type-II alveolar epithelial cell line designated T7, derived from transgenic H-2K(b)-tsA58 mice, capable of being passaged as an immortalized cloned cell line in culture. H-2K(b)-tsA58 mice harbor a temperature-sensitive (ts) mutant of the simian virus 40 (SV40) large tumor antigen (T antigen) under the control of the gamma-interferon (INF)-inducible mouse major histocompatibility complex H-2Kb promoter. When cultured under permissive conditions (33 degrees C and in the presence of gamma-INF) cells isolated from H-2Kb-tsA58 mice express the large T antigen, which drives the cells to proliferate. However, upon withdrawal of the gamma-INF and transfer of the cells to a higher temperature (39 degrees C), T antigen expression is turned off, the cells stop proliferating and differentiate. The T7 cell line is a clonal cell line originally derived from a Type-II cell-rich fraction isolated from lungs of H-2Kb-tsA58 mice. The T7 cells form confluent monolayers, and have a polarized epithelial cell morphology with tight junctions and apical microvilli. In addition, the T7 cells have distinct cytoplasmic lamellar bodies, which become more numerous and pronounced when the cells are grown under nonpermissive conditions. The T7 cells synthesize and secrete phosphatidylcholine and the three surfactant proteins, SP-A, SP-B, and SP-C. The T7 cell line is unique in that it is the first non-tumor-derived Type-II cell line capable of synthesizing and secreting the major components of surfactant. Based on the criteria studied, the T7 cell line is phenotypically very similar to normal Type-II cells. The T7 cell line, therefore, should prove a valuable experimental system to advance the study of the cell biology/physiology of surfactant metabolism and secretion as well as serve as a model for other studies of Type-II cell physiology.

Embryonic and early fetal development of human lung vasculature and its functional implications.

Recently, we have identified in the mouse three processes involved in the early development of pulmonary vasculature: angiogenesis for branching of central vessels, vasculogenesis (lakes in the mesenchyme) for peripheral vessels, and a lytic process to establish luminal connection between the two. We have established that these three processes also operate in the human by studying serial sections of human embryos and early fetuses. Vascular lakes of hematopoietic cells appear at stage 13, i.e., 4+ weeks gestational age (GA), the first intrapulmonary vascular structure to appear. At stage 20 (50.5 days GA), a venous network with luminal connections to central pulmonary veins (PV) is present. Airways have not yet reached these regions of lung. At its first intrapulmonary appearance, the pulmonary artery (PA) is small and thick walled: it runs with the airway but its branching is slower, so many peripheral airways are not accompanied by a PA branch. By contrast, the PV has a peripheral patent network well before the PA. In the pseudoglandular phase, airway branching continues, and the PA catches up so that small PA branches are found with all airways. Later in this phase small nonmuscular vessels lie in the mesenchyme close to airway epithelium. By the early canalicular phase and the age of viability, continuity between pulmonary artery and the peripheral capillary network must be established. In a 10-week fetus several structures suggesting a breakthrough site were seen. Air-blood barrier structure is first seen at 19 weeks. Thus in the lung, the PA and PV are dissociated in their timing and pattern of branching. Early veins are present diffusely through the mesenchyme and establish central luminal connection to the main pulmonary vein before airway or artery are present at this level.

An alternatively spliced surfactant protein B mRNA in normal human lung: disease implication.

We identified an alternatively-spliced surfactant protein B (SP-B) mRNA from normal human lung with a 12 nt deletion at the beginning of exon 8. This deletion causes a loss of four amino acids in the SP-B precursor protein. Sequence comparison of the 3' splice sites reveals only one difference in the frequency of U/C in the 11 predominantly-pyrimidine nucleotide tract, 73% for the normal and 45% for the alternatively-spliced SP-B mRNA (77-99% for the consensus sequence). Analysis of SP-B mRNA in lung indicates that the abundance of the alternatively-spliced form is very low and varies among individuals. Although the relative abundance of the deletion form of SP-B mRNA remains constant among normal lungs, it is found with relatively higher abundance in the lungs of some individuals with diseases such as congenital alveolar proteinosis, respiratory distress syndrome, bronchopulmonary dysplasia, alveolar capillary dysplasia and hypophosphatasia. This observation points to the possibility that the alternative splicing is a potential regulatory mechanism of SP-B and may play a role in the pathogenesis of disease under certain circumstances.

Spontaneous resolution of myelodysplastic cytogenetic abnormality developed during the treatment of leukemia.

PURPOSE: We describe the spontaneous resolution of a myelodysplastic cytogenetic abnormality developing during the treatment of acute lymphocytic leukemia. PATIENTS AND METHODS: A 6-year-old girl with acute lymphocytic leukemia had a clinical picture of myelodysplasia 18 months after diagnosis. The clonal cytogenetic abnormality, 46,XX,del(5)(q12q12), resolved spontaneously 4 months after the discontinuation of chemotherapy. Maintenance chemotherapy was resumed 1 month later and continued for an additional 9 months. Currently, she has been off therapy for 10 months. CONCLUSION: A myelodysplastic clonal cytogenetic abnormality developing during treatment may cause some confusion for management. This study demonstrates that spontaneous resolution is possible and that bone marrow transplantation or other intensive treatment may not be necessary.

The effect of intrauterine growth restriction upon fetal and postnatal hepatic glucose transporter and glucokinase proteins.

Employing immunohistochemical and Western blot analyses, we investigated the cellular localization (22-d fetal and 14-d postnatal animals) and concentrations (22-d fetal to 21-d postnatal animals) of rat hepatic glucose transporters (Glut 1 and Glut 2) and glucokinase in response to development and uteroplacental insufficiency with IUGR. Glut 1, the predominant fetal hematopoietic cellular isoform, persisted in postnatal hematopoietic islands and was noted minimally in fetal hepatic cellular membranes. A approximately 40% extrauterine decline in Glut 1 levels paralleled the decline in hematopoietic cells. IUGR increased the fetal hepatic Glut 1 levels in parallel with an expanded hematopoietic cell mass (p < 0.05). In contrast, IUGR failed to alter the 2-fold increase in extrauterine Glut 2 concentrations (1-7-d postnatal animals), the isoform found in fetal and postnatal hepatocytic cell membranes. Glucokinase, the nuclear enzyme, increased 25% postnatally. IUGR caused a 16% increase in fetal glucokinase levels and a approximately 25% decline at postnatal d 1 (p < 0.05) without a comparable change in the hepatocytic cell number (92 +/- 6 versus 86 +/- 4). We conclude that hepatic Glut 1 concentrations reflect the extramedullary hematopoietic cellular mass, whereas extrauterine Glut 2 changes herald the need for enhanced flexibility in hepatocytic glucose transport with the initiation of food ingestion. The age-related alteration along with the IUGR-induced compensatory changes in the nuclear-mitochondrial glucokinase levels attributes a critical role for this enzyme in perinatal hepatocytic glucose homeostasis.

Partial deficiency of surfactant protein B in an infant with chronic lung disease.

OBJECTIVE: To evaluate components of pulmonary surfactant and identify mutations in the surfactant protein B gene (SP-B) of a term infant with severe respiratory distress and chronic lung disease. PATIENT AND TESTING: Respiratory distress developed in an infant delivered at term, and he required extracorporeal bypass support for 2 weeks. Until his unexpected death at 9.5 months, he was ventilator and oxygen dependent and required continual dexamethasone therapy. Tracheobronchial lavage samples were analyzed for content of surfactant proteins (SPs), and DNA from blood samples were sequenced and analyzed by polymerase chain reaction restriction analysis for the presence of SP-B gene mutations. Surfactant lipid composition and function, the contents of SPs and their messenger RNAs (mRNAs), and the immunostaining pattern for SPs were determined in postmortem lung tissue. RESULTS: The lavage sample contained SP-A but not SP-B, and DNA restriction analysis indicated that the patient and his mother were heterozygous for the previously described 121ins2 mutation of SP-B. Postmortem lung tissue contained normal levels of SP-A and its mRNA, a low but detectable level of SP-B, and near normal content of SP-B mRNA. SP-C was abundant on staining, and some 6-kd precursor was present in tissue. A surfactant fraction was deficient in phosphatidylglycerol and was not surface active. On DNA sequencing, a point mutation was found in exon 7 of the patient's SP-B gene allele without the 121ins2 mutation, resulting in a cysteine for arginine substitution, and the father was a carrier for the same mutation. CONCLUSIONS: We describe a patient who is a compound heterozygote with a new mutation and only a partial deficiency of SP-B. Some forms of inherited SP-B deficiency may have low expression of immunoreactive and possibly functional SP-B with milder lung disease and longer survival. These infants may benefit from glucocorticoid therapy and may not develop antibodies to SP-B after either lung transplant or gene therapy.

Surfactant protein B deficiency: antenatal diagnosis and prospective treatment with surfactant replacement.

An infant with a family history of congenital alveolar proteinosis associated with surfactant protein B (SP-B) deficiency was identified when SP-B was not detected in amniotic fluid obtained at 37, 38, and 40 weeks of gestation. Surfactant replacement with commercially available preparations that contained SP-B was begun soon after delivery. Progressive respiratory failure developed despite continued surfactant replacement, corticosteroid therapy, and extracorporeal membrane oxygenation. The infant died at 54 days of age while awaiting lung transplantation. Surfactant extracted from amniotic fluid, bronchoalveolar lavage fluid, and lung tissue had no phosphatidylglycerol; surface tension was 24 dynes/cm (normal, < 10 dynes/cm) and did not decrease with in vitro addition of exogenous SP-B. Pulmonary vascular permeability measured with positron emission tomography was twice normal. At autopsy the alveolar proteinosis pattern was less prominent than that seen in affected siblings. Immunoreactivity of SP-B was absent in type II cells, but numerous foreign body granulomas with central immunoreactivity for SP-B and surfactant protein C were present. We conclude that exogenous surfactant replacement did not normalize surfactant composition, activity, or pulmonary vascular permeability. These findings suggest that endogenous SP-B synthesis is necessary for mature surfactant metabolism and function.

Ultrastructure of lung in surfactant protein B deficiency.

Congenital alveolar proteinosis (CAP), a cause of respiratory failure in fill-term newborns, often leads to death in infancy despite medical therapy. We recently described an inherited deficiency of surfactant protein B (SP-B) (N. Engl. J. Med. 1993; 328:406-410) in two siblings with CAP. The SP-B deficiency was accompanied by marked abnormalities, both quantitative (increase) and qualitative (distribution), of SP-A and SP-C in the lungs of the affected infants. Ultrastructural studies of the lung of one of these infants and of a third affected sibling born in the index family showed abundant alveolar concentric multilamellated structures and membranous vesicles but no typical tubular myelin. In addition, membranous vesicles from type II cells and immunogold labeled SP-A and SP-C were found between type II cells and their basement membrane despite intact interepithelial cell junctions. These findings suggest an important role for SP-B in the directionality of surfactant secretion and in the formation of tubular myelin.

Hyperoxia induces interstitial (type I) and increases type IV collagenase mRNA expression and increases type I and IV collagenolytic activity in newborn rat lung.

Oxygen toxicity is attributed to the reaction of oxygen metabolites with cellular components leading to cell destruction. Activation of latent human neutrophil interstitial collagenase by reactive oxygen species has been demonstrated. The potential role of collagenases in hyperoxic lung injury has not been investigated. We studied the effect of hyperoxia on newborn rat lung water content, morphology and ultrastructure, interstitial (type I) and type IV collagenase gene expression and type I and IV collagenolytic activity. We observed that hyperoxia causes pulmonary edema, alters newborn rat lung morphology in a sequential manner and produces ultrastructural alterations, induces type I and increases type IV collagenase mRNA expression, and increases type I and IV collagenolytic activity. A role for type I and IV collagenase in hyperoxic newborn lung injury or in the recovery following the injury is proposed.

Effects of indomethacin in utero on the pulmonary vasculature of the newborn guinea pig.

The clinical syndrome of persistent pulmonary hypertension of the newborn results from failure of the normal perinatal vascular adaptation, and functionally is characterized by persistent right to left shunting of blood through the foramen ovale and ductus arteriosus. Exposure of the fetus to drugs that inhibit prostaglandin synthesis and cause closure of the ductus arteriosus has been suggested as one cause of persistent pulmonary hypertension of the newborn. We attempted to produce a functional and structural model of persistent pulmonary hypertension of the newborn by administration of indomethacin, a cyclooxygenase inhibitor, to pregnant guinea pigs. Five pregnant guinea pigs received 3.5 mg/kg indomethacin intravenously twice each day for the 12 to 19 days before delivery and seven controls received saline. Hemodynamic studies were performed in eight "treated" newborns and in 12 controls. After sacrifice, the ductus was ligated and, for morphometric studies, the pulmonary arteries were distended with barium/gelatin. The treated animals did not show the intraacinar structural or hemodynamic changes of persistent pulmonary hypertension of the newborn. It seems that the indomethacin did cross the placenta because lung structure was modified. The radical alveolar count and alveolar/artery ratio were increased and the preacinar arteries dilated, with more increase in muscle mass. This could be explained by increased pulmonary blood flow because of ductal constriction but direct effect of indomethacin cannot be excluded.

Myofibrillar degeneration and necrosis of the visceral smooth musculature: an ischemic visceral myopathy.

Pathologic studies of the visceral smooth musculature in humans are scant despite the relatively frequent occurrence of alterations in these muscles in autopsy material. We investigated the different types of lesions of this musculature observed in various conditions associated with ischemia--acute tubular necrosis, congenital heart disease (low output syndrome due to open heart surgery), and necrotizing enterocolitis in premature babies. Control cases included normal rat tissue undergoing autolysis and rigor mortis and bowel resected from patients with ulcerative colitis and Hirschsprung's disease. Four histologically distinct lesions were present on hematoxylin--eosin staining in the ischemic group: contraction bands, wavy fibers, thick waves, and coagulation necrosis. These lesions were absent in the control groups. We conclude that myofibrillar degeneration and necrosis of the visceral musculature are common in disorders associated with visceral ischemia. These changes are not artifacts produced by autolysis, rigor mortis, or technical handling, nor are they induced by nonischemic inflammatory conditions. Catecholamines may play a role in their genesis.

Midline cervical cysts in children. Thyroglossal anomalies.

Deep, midline cervical cysts clinically diagnosed as thyroglossal duct cysts (TDCs), have been pathologically classified as dermoid cysts because of the presence of skin appendages and a squamous epithelial lining. In 75 midline cervical masses preoperatively diagnosed as TDC, we could classify only 54 as TDC, using the preexisting criteria of squamous or ciliated columnar epithelial lining associated with a tract or thyroid follicles. Eleven cysts were reclassified as dermoid, and six were called "mixed" because of features of both dermoid cysts (skin appendages) and TDC (epithelial tract or thyroid follicles). The morphological similarity of all these lesions suggests a common origin, perhaps from totipotential tissue entrapped during the descent of the embryonic thyroglossal duct from the base of the tongue. We conclude that these lesions should be grouped together under the eponym of "thyroglossal anomalies," and that treatment for all should consist of the Sistrunk procedure.

Recurring digital fibroma of childhood.

A recurrent digital fibroma of childhood is reported. This case illustrates difficulties in the management of these recurrent tumors. Despite tumor-free margins on the excised tumors, recurrence occurred at other sites. Recurrences or new primary lesions are reported in 75% of the cases. Because of the rare tendency of these lesions to regress and the high recurrence rate, an individualized approach based on lesion location and behavior is recommended.