Fetal Cells in the Murine Maternal Lung Have Well-Defined Characteristics and Are Preferentially Located in Alveolar Septum (original) (raw)
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Chimerism
During pregnancy, fetal cells cross into the maternal organs where they reside postpartum. Evidence from multiple laboratories suggests that these microchimeric fetal cells contribute to maternal tissue repair after injury. In mouse models, most injury experiments are performed during pregnancy; however, in a clinical setting most injuries or diseases occur postpartum. Therefore, experiments using animal models should be designed to address questions in the time period following delivery. In order to provide a baseline for such experiments, we analyzed the natural history of fetal cells in the postpartum maternal organs. Female C57BL/6J mice were mated to males homozygous for the enhanced green fluorescent protein gene. Fetal cells in the maternal lungs and bone marrow were identified by their green fluorescence using in a high-speed flow cytometer and their counts were compared between the lung and bone marrow. Spearman correlation analysis was used to identify relationships betwee...
Biology of Reproduction, 2008
The purpose of the study was to understand in more detail the natural history of fetomaternal cell trafficking in healthy pregnant mice. Our goal was to identify the best target organs and days during pregnancy for further mechanistic studies of the role of fetal cells in maternal disease and injury. C57BL/6J wildtype virgin females (n ¼ 54) were mated with congenic enhanced green fluorescent protein (EGFP) transgenic males. During pregnancy and after delivery, female mice were euthanized, and eight organs and blood were analyzed for the presence of fetal GFP þ cells with flow cytometry and real-time quantitative PCR. Maternal lungs, liver, and spleen were also analyzed by fluorescent stereomicroscopy. Fetal GFP þ cells were first found at low frequency at Embryonic Day 11, increased to a maximum at Embryonic Day 19, and decreased rapidly postpartum. These fetal cell dynamics were significantly reproducible among all mice studied. In addition, there was a consistent distribution of fetal cells within maternal organs, with lung, liver, blood, and spleen having the greatest concentrations; these were highly correlated at all time points (P , 0.0001). Maternal lung contained 10-to 100-fold more fetal cells than any other organ, and using all three techniques, the number of fetal cells detected was the most consistent and reproducible in this organ. Stereomicroscopy showed that within the lung, fetal cells were widely and apparently randomly distributed. Using a murine model, our data demonstrate that fetomaternal cellular trafficking occurs in reproducible patterns, is maximal near term delivery, and has predilection for the maternal lung.
Development of human fetal lung in organ culture compared with in utero ontogeny
In Vitro Cellular & Developmental Biology - Animal, 1993
In utero, at around 23 wk gestation, the progenitor epithelium of distal airway differentiates into type I and type II pneumatocytes. Human fetal lung organ cultures, as early as 12 wk gestation, have the competence to self-differentiate. Distal airway epithelial immunoreactivity to cytokeratins CK 7, 8, and 18 decreases with differentiation both in utero and in organ culture, whereas reactivity to epithelial membrane antigen remains constant in both. As distal airways dilate, the mean percentage airspace of fetal lungs in organ culture increases to 58%, equivalent to lung of gestation 26.0 + 7.3 wk. In organ culture, capillary blood vessels, visualized by vimentin immunoreactivity, remodel and more closely approximate the epithelium but without direct invasion. In utero, at 23 wk gestation, elastin appears as condensation around airways and forms a basis for secondary crests which, by 29 wk gestation, evolve into alveolar septae. In organ culture, no elastin is deposited, no secondary or alveolar crests form, and the lung retains a simple saccular structure. Differentiation of the terminal airway epithelium and mesodermal maturational events to facilitate gas exchange, such as capillary invasion or secondary-alveolar crest formation, are almost synchronous in human lung in utero but clearly dissociate in organ culture.
The Anatomical record, 1992
To evaluate further the role of type II alveolar epithelial cells in primate lung development, lungs of fetal (46 to 155 days gestational age [DGA]), postnatal, and adult rhesus monkeys were investigated with antibodies against surfactant protein A (SP-A), Alcian blue (AB) staining, and periodic acid-Schiff (PAS) staining with/without alpha-amylase pre-treatment. In adult and postnatal lungs, type II cells (cuboid shape; large, roundish nucleus) displayed a unique cytoplasmic staining for SP-A. In prenatal lungs, a low-columnar to cuboid type of cell with a large, roundish nucleus was first detectable by 62 DGA. It was the only cell type to line the distalmost tubules or buds of the prospective respiratory tract. It exhibited (initially partial) cytoplasmic staining for SP-A. AB and PAS stainings showed the presence of acid glycoconjugates and large apical and/or basal glycogen fields. After 95 DGA, the lining of the distal respiratory tract additionally displayed flatter cells with...
Microscopy Research and Technique, 1993
The aim of this study was to describe and compare the ultrastructural features and functional maturity of alveolar epithelial cells in hypoplastic and normal fetal rat lungs. Pulmonary hypoplasia in association with congenital diaphragmatic hernia was induced in fetuses by administration of 2,4-dichlorophenyl-p-nitrophenylether (Nitrofen) to pregnant Sprague Dawley rats (100 mg on day 10 of gestation). Lung tissue of Nitrofen-exposed and control fetal rats aged 19-22 days (vaginal plug day 1, birth day 23) was embedded in Epon. Semithin (1 km) toluidine blue-stained sections were examined by light microscopy; ultrathin sections (-80 nm) were studied via transmission electron microscopy. In bronchoalveolar lavage fluid from control and Nitrofenexposed fetuses (day 221, phospholipid fractions and surfactant protein A content were measured semiquantitatively. On day 19 both control and Nitrofen-exposed lungs contained only cuboid alveolar epithelial cells; from day 20 there were cuboid, low cuboid, and thinner epithelial cells. The (low) cuboid cells contained large glycogen fields, some precursory stages of multilamellar bodies (MLBs), and just a few mature MLBs on day 19 and 20; smaller glycogen fields, more precursory stages, and more mature MLBs on day 21; and little or no glycogen but many precursory stages and mature MLBs on day 22. The thinner cells contained little or no glycogen and a few precursory stages of MLBs on days 20-22; very thin cells on day 22 contained neither glycogen nor any precursory stages of MLBs. MLBs and tubular myelin were seen in the lumens of future air spaces from day 20 onward. Nitrofen-exposed lungs differed from control lungs in that inclusion bodies (IBs) were less numerous in (low) cuboid alveolar cells on days 19 and 20, and more glycogen was seen on day 22. In addition intra-and extracellular "MLBs" in exposed lungs more often had a n unusual appearance, i.e., a confluent structure and higher electron density. However, despite morphologic differences, there was no clear difference in phospholipid composition and SP-A content per mol phospholipid in bronchoalveolar lavage fluid. We conclude that morphologically hypoplastic lungs are less mature near term, without an apparent effect on surfactant composition. 0 1993 Wiley-Liss, Inc.
Differentiation of xenografted human fetal lung parenchyma
Early Human Development, 2008
The goal of this study was to characterize xenografted human fetal lung tissue with respect to developmental stage-specific cytodifferentiation. Human fetal lung tissue (pseudoglandular stage) was grafted either beneath the renal capsule or the skin of athymic mice (NCr-nu). Tissues were analyzed from 3 to 42 days post-engraftment for morphological alterations by light and electron microscopy (EM), and for surfactant protein mRNA and protein by reverse transcription-polymerase chain reaction (RT-PCR) and immunocytochemistry (ICC), respectively. The changes observed resemble those seen in human lung development in utero in many respects, including the differentiation of epithelium to the saccular stage. Each stage occurred over approximately one week in the graft in contrast to the eight weeks of normal in utero development. At all time points examined, all four surfactant proteins (SP-A, SP-B, SP-C, and SP-D) were detected in the epithelium by ICC. Lamellar bodies were first identified by EM in 14 day xenografts. By day 21, a significant increase in lamellar body expression was observed. Cellular proliferation, as marked by PCNA ICC and elastic fiber deposition resembled those of canalicular and saccular in utero development. This model in which xenografted lung tissue in different stages of development is available may facilitate the study of human fetal lung development and the impact of various pharmacological agents on this process.
Culture of differentiated and undifferentiated type II cells from fetal rat lung
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 1987
We have developed a relatively simple and reproducible method for the isolation and culture of both differentiated and undifferentiated type II cells from fetal rat lung. The technique involves an initial period of explant culture in serum and hormone free medium, followed by enzymatic dissociation of the explants, differential adhesion to remove fibroblasts, incubation of the cell pellet to promote aggregation of the type II cells and monolayer culture of the type |I cells. The type II cells form clusters which are surrounded by scattered fibroblasts. When the technique was performed with three differential adhesion steps, cultures contained 86.0 + 1.4% type II cells. To obtain a higher degree of purity and greater yield, two differential adhesions followed by gentle trypsinization of the cultures which selectively removes the isolated fibroblasts was performed. This resulted in cultures with 89.4 + 1.7% type II cells. The differentiated fetal type II cell cultures were prepared from 19-day fetal rat lungs which were initially maintained in explant culture for 48 h. These differentiated cells demonstrated the characteristic morphologic features of type II cells including lamellar bodies and microvUli. Undifferentiated fetal cells were prepared in a similar manner from 18-day fetal rat lung maintained in explant culture for 24 h. These cells did not contain intracellular osmiophilic granules; the appearance of these granules could, however, be induced by hormones. For this reason they are considered to be pre-type II cells. The viability of the cultured cells was 97%. Both the differentiated and undifferentiated fetal type II cells specifically bound the Maclura pomifera lectin, a type II cell surface marker. The phospholipid profile of the fetal cells was similar to that of adult rat type II cells; the differentiated fetal cells, however, synthesized less phosphatidylcholine than the adult cells did, but more than the undifferentiated fetal cells. The differentiated fetal cells secreted phosphatidylcholine at a basal rate of 0.6% ± 0.1% during a 90-min incubation. There was dose-dependent stimulation of phosphatidyicholine secretion after exposure to terbutaline. Maximum stimulation (76%) was observed at a concentration of 10 /~M. This culture system provides a valuable model for studies of the maturation of the undifferentiated fetal type II cell and surfactant metabolism and secretion in the differentiated fetal type |I cell.
Microchimeric fetal cells cluster at sites of tissue injury in lung decades after pregnancy
Reproductive BioMedicine Online, 2008
Fetal cells trafficking into maternal blood during pregnancy engraft tissues and persist for decades in marrow and bone. While persistent fetal cells were initially implicated in autoimmune disease, animal studies suggest that microchimeric fetal cells play a broader role in response to tissue injury. This study investigated whether fetal cells participate in tissue repair after human pregnancy. Specimens were obtained from women undergoing surgery for suspected lung cancer. Y-fluorescence in-situ hybridization was performed on paraffin-embedded sections, with the investigator blinded to medical histories. Male cells were identified in lung/thymus tissue from all women with known male pregnancies, but not in those without sons. The frequency of male microchimeric cells was seven-fold greater in lung/thymus tissues than marrow and was two-fold greater than normal bone from the same women. Nested-polymerase chain reaction for sex determining region Y confirmed male DNA in tissues. Male cells in lung were clustered in tumour rather than surrounding healthy tissues. In conclusion, male presumed-fetal cells were identified in pathological post-reproductive tissues, where they were more likely to be located in diseased tissues at several-fold higher frequency than normal tissues. It is suggested that fetal cells are present at sites of tissue injury and may be stem cells, either recruited from marrow or having proliferated locally.