Lung tissues of rats from two different age groups (2-12 and 16-26 months of age) were studied by both light and electron microscopy. Proliferation of granular pneumocytes in pulmonary alveolar lining was a frequent occurrence in older rats. Lungs of older rats showed not only an increase in number of granular pneumocytes, but also a remarkable increase of lamellar bodies and other forms of lipid vacuoles in individual granular pneumocytes. Spontaneously-occurring nodular lesions characterized by the accumulation of macrophages in the alveolar spaces were accompanied by desquamation and proliferation of granular pneumocytes. These lesions developed only in the lungs of rats older than l7 months of age. Such lessions in lungs of old rats were similar in many respects to desquamative interstitial pneumonitis of human lungs. Atrophy of alveolar walls and emphysematous areas seen in senile rats was characterized by irregular cytoplasmic breakdown of Type I alveolar lining epithelial cells. Obliteration of capillaries by spontaneously-occurring thrombus formation or a herniated cytoplasm of the septal cell and collagen fibers was considered to be a cause of atrophy of alveolar walk. Degeneration and actual breakdown of endothe1ial cytoplasm of puImonary capillaries enhanced herniation of the septal tissue. Vacuolar degeneration of epithelial cytoplasm was occasionally observed, but only in rats older than 20 months of age. The basement membrane of pulmonary alveolar walls was often thicker in old rats than in younger rats. Hyperplasia of granular pneumocytes invariably accompanied large septal cells, some of which contained many of the organelles found in granular pneumocytes.
Aging* ; Animal ; Female ; Macrophages/cytology* ; Male ; Pulmonary Alveoli/anatomy & histology* ; Pulmonary Alveoli/cytology ; Rats

To establish a better method of primary culture for alveolar epithelial type II cells (AEC II) and to study its bionomics, alveolar epithelial type II cells were isolated by digestion with trypsin and collagenase, which were then purified by plated into culture flask coated with rat immunoglobulin G. The purified AEC II were identified by alkaline phosphatase staining, electron microscopy, immunocytochemical staining of pulmonary surfactant protein A (SPA). The SPA expression and transfection characteristics were compared with those of A549 cell line. The results showed that AEC II could be isolated by digestion with trysin and collagenase and purified by adhesive purification by using IgG, with a yield of about 2-3 x 10(7), and a purity of about 75%-84%. Cells could be quickly identified with AKP staining. AEC II were different from A549 cell line in terms of SPA expression and transfection characteristics. It is concluded that adhesive purification with IgG can improve the purity of AEC II, and AKP staining is simple in cell identification. AEC II can not be completely replaced by A549 cells in some studies because the differences between them, such as SPA expression.
Cell Culture Techniques/*methods ; Cell Separation/*methods ; Cells, Cultured ; Ecology ; Epithelial Cells/*cytology ; Immunoglobulin G/pharmacology ; Pulmonary Alveoli/*cytology ; Pulmonary Surfactant-Associated Protein A/biosynthesis ; Rats, Wistar

OBJECTIVETo investigate the expression patterns of surfactant protein B (SP-B) and its role in the development of human fatal lung epithelial cells.

METHODSHuman fetal lung tissues were obtained from 37 fetuses of 10-34 weeks at abortion with parental consent and from two newborn infants who died of non-pulmonary causes. SP-B expression in the lung tissues was examined by immunohistochemistry.

RESULTSSP-B was detected in the cytoplasm of nonciliated columnar epithelial cells of the human fetal lung in as early as the 16th week of gestation. The positive reaction of SP-B was enhanced during canalicular stages and was more intense in the distal than in the proximal airway epithelium. From the 25th week to the prenatal stage, SP-B expression underwent no significant changes in the primitive alveolar stage, but increased remarkably after birth.

CONCLUSIONThe expression and secretion of SP-B reflects the maturation of the epithelial cells in human fatal lungs, and may closely associate with the survival ability of the newborn infants.

Cell Survival ; physiology ; Cells, Cultured ; Epithelial Cells ; cytology ; metabolism ; Fetus ; Humans ; Infant, Newborn ; Lung ; Pulmonary Alveoli ; cytology ; metabolism ; Pulmonary Surfactant-Associated Protein B ; biosynthesis ; physiology

To study the difference of changes on apoptosis, necrosis and respiratory burst of the polymorphonuclear neutrophils (PMN) in endotoxemia rat model. LPS (O(55)B(5), 5 mg/kg) was injected into abdominal cavity of 20 random normal Wistar rat. 2, 4, 8 and 12 hours after injection, the changes of apoptosis, necrosis and respiratory burst of the rats between PMN from the peripheral blood and from the bronchoalveolar lavage fluid were observed using the flow cytometer. At the same time, 5 uninjected rats were taken as control. The results demonstrated that the quantity proportions of apoptosis of PMN between the peripheral blood PMN and the bronchoalveolar lavage fluid PMN in rat's endotoxemia were similar. However, comparison with the uninjected LPS rat, the necrosis of peripheral blood PMN obviously increased and the respiratory burst capacity was clearly inhibited. Contrarily, the necrosis of bronchoalveolar lavage fluid PMN obviously decreased and the respiratory burst obviously increased in the injecting LPS rat. It was concluded that the necrosis and apoptosis displayed differently between the pulmonary and peripheral blood PMNs in endotoxemia. Under state of inflammation, the surviving PMN in tissue increased and kept the activated state due to tissue injury.
Animals ; Apoptosis ; Bronchoalveolar Lavage Fluid ; cytology ; Endotoxemia ; blood ; Necrosis ; Neutrophils ; physiology ; Pulmonary Alveoli ; pathology ; Rats ; Rats, Wistar ; Respiratory Burst

OBJECTIVETo observe the homing and differentiation of marrow-derived mesenchymal stem cells (MSC) transplanted intravenously in smoke inhalation injured rabbits.

METHODSThirty-two New Zealand big ear rabbits were divided into normal control group (NC), inhalation injury group (II), normal control + MSC treatment group (NM), and MSC treatment group (MT) according to the random number table, with 8 rabbits in each group. Rabbits in NC group were injected with 10 mL phosphate buffered saline (PBS) via ear marginal vein. Rabbits in NM group were injected with 10 mL PBS containing the third generation MSC labeled by BrdU (1 × 10(7) per 10 mL PBS) via ear marginal vein. Severe smoke inhalation injury model was reproduced in the other two groups, among them rabbits in II group were treated as rabbits in NC group, rabbits in MT group treated as rabbits in NM group. On the 7th and 28th day post treatment (PTD), lung tissue and trachea tissue were harvested from four groups for observation on injury with HE staining. Homing of MSC in injured tissue was observed with immunohistochemistry staining. The differentiation of MSC into functional cells was observed with immunohistochemical double staining of combining nuclear marker BrdU with lung (trachea) membrane-specific marker aquaporin-5 (AQP-5), alkaline phosphatase (AKP), CD34, and cytokeratin respectively.

RESULTS(1) MSC homing in lung and trachea tissue was observed in MT group on PTD 7, which was not observed in NM group. (2) AQP-5, AKP, and CD34 positive MSC were observed in lung tissue in MT group on PTD 28, while cytokeratin positive MSC was not observed in trachea tissue. No positively marked MSC was observed in NM group. (3) Injury in lung and trachea was less severe in MT group than in II group; and the proliferation of fibroblasts was less in MT group.

CONCLUSIONSIntravenous injection of MSC to rabbits with smoke inhalation injury can migrate to lung and trachea tissue at obviously inflammatory site, and differentiate into alveolar epithelial cells typeI and II, and pulmonary vascular endothelial cells, which may participate in the process of tissue repair in smoke inhalation injury.

Animals ; Bone Marrow Cells ; cytology ; Cell Differentiation ; Cells, Cultured ; Endothelial Cells ; cytology ; Epithelial Cells ; cytology ; Lung ; cytology ; Mesenchymal Stem Cell Transplantation ; Mesenchymal Stromal Cells ; cytology ; Pulmonary Alveoli ; cytology ; Rabbits ; Smoke Inhalation Injury ; pathology ; Trachea ; cytology

OBJECTIVETo describe the pathologic features and diagnostic algorithm of pulmonary alveolar proteinosis (PAP).

METHODSThirty-nine biopsy and postmortem cases of PAP were studied by light microscopy and histochemical staining using periodic acid-Schiff (with digestion) (PAS-D), mucicarmine (with digestion) (mucicarmine-D) and alcian blue.

RESULTSHistologically, the affected lung tissue displayed the following characteristic features: (1) alveoli and some of the small bronchioles were filled with eosinophilic and fine granular proteinaceous material with needle-like clefts; (2) proteinaceous material was seen admixed with various numbers of degenerated and sometimes exfoliated pneumocytes; (3) pneumocytes were hyperplastic; (4) alveolar capillaries and alveolar septa had become hyperemic, but pulmonary interstitial inflammation was not obvious; (5) no significant inflammation was identified in the bronchial wall; (6) compensating emphysema was noted in the surrounding lung parenchyma. Fragments of eosinophilic, finely granular proteinaceous material with needle-like clefts were also found in the bronchoalveolar lavage fluid under light microscopy. The proteinaceous material was stained red by PAS-D. The staining for mucicarmine-D was negative, while alcian blue staining was either weakly positive (faint blue staining) or negative. Pathologic examination of lung biopsies and bronchoalveolar lavage fluid thus remaines the gold standard for diagnosis of PAP.

CONCLUSIONSIdentification of homogeneous, eosinophilic, finely granular and PAS-D-positive proteinaceous material with needle-like clefts in alveolar spaces or bronchoalveolar lavage fluid is of diagnostic importance in PAP. Bronchoalveolar lavage, being a relatively safe and non-invasive procedure, can be a useful adjunct in arriving at the final conclusion.

Adult ; Bronchoalveolar Lavage ; Bronchoalveolar Lavage Fluid ; cytology ; Female ; Humans ; Lung ; pathology ; Male ; Middle Aged ; Periodic Acid-Schiff Reaction ; Pulmonary Alveolar Proteinosis ; pathology ; therapy ; Pulmonary Alveoli ; pathology

OBJECTIVETo investigate the effect of cryptoporus polysaccharide(CP)on lipopolysaccharide(LPS)-induced production of monocyte chemoattractant protein-1(MCP-1)in human lung epithelial A549 cells.

METHODSA549 cells were stimulated with LPS in the presence or absence of CP. The protein concentration and mRNA expression of MCP-1 were determined by enzyme-linked-immunosobent assay(ELISA)and semi-quantitative RT-PCR, respectively.

RESULTThe protein concentration of MCP-1 was significantly increased by LPS 1000 microg/L at 24 h. There were no effects on the growth and viability of A549 cells in the presence of CP 100 microg/L or dexamethasone 1 mumol/L. However, CP 100 microg/L or dexamethasone 1 micromol/L significantly inhibited the protein concentration and mRNA expression of MCP-1 induced by LPS.

CONCLUSIONCP can regulate MCP-1 production, which may be associated with its effects on lung inflammation.

Cell Line ; Chemokine CCL2 ; genetics ; metabolism ; Epithelial Cells ; cytology ; metabolism ; Humans ; Lipopolysaccharides ; pharmacology ; Polyporaceae ; chemistry ; Polysaccharides ; isolation & purification ; pharmacology ; Pulmonary Alveoli ; cytology ; RNA, Messenger ; metabolism

OBJECTIVETo investigate the effects of cysteinyl leukotriene (CysLT) receptor agonist leukotriene D4 (LTD4) on proliferation and migration in lung epithelial A549 cells.

METHODSThe expression of CysLT1 receptor and CysLT2 receptor was determined by immunofluoresence staining in A549 cells. A549 cells were treated with LTD4 (0.01-100 nmol/L) for 24-72 h. Cell viability was detected by MTT reduction assay. Cell migration was determined by modified scratch and healing model.

RESULTSIn A549 cells, CysLT1 receptor and CysLT2 receptor were mainly expressed in the cytoplasm, membrane and few in the nuclei. The treatment of LTD4 (0.01-100 nmol/L) for 24-72 h caused no effect on cell viability (Ps>0.05); when A549 cells were treated with 100 nmol/L LTD4 for 24, 48 and 72 h the cell viability was (103.00±4.46)%，(107.00±9.45)% and (105.00±9.02)% of control, respectively (Ps>0.05). The migration rate of A549 cells after scratching during the first 24 h was markedly greater than that during the second and third 24 h in the same concentration groups; however, no significant difference in migration rate was noticed when the cells were treated with different concentrations of LTD4 (0.01-100 nmol/L)(Ps>0.05). The migration of A549 cells was 1.15-fold, 1.21-fold and 1.06-fold of that of control when the cells were treated with 100 nmol/L LTD4 for 24, 48 and 72 h, respectively (Ps>0.05).

CONCLUSIONThe proliferation and migration of A549 cells are not changed when treated with 0.01-100 nmol LTD4 for up to 72h.

Cell Line ; Cell Movement ; drug effects ; Cell Proliferation ; drug effects ; Epithelial Cells ; cytology ; drug effects ; Humans ; Leukotriene D4 ; pharmacology ; Pulmonary Alveoli ; cytology

Acute respiratory distress syndrome (ARDS) remains a poor prognosis in spite of the recent development of new therapeutic strategies. Cell-based therapy with stem cells has been considered as a promising way for the treatment of vital organ damage. Putative endogenous stem cells have been shown to be located within the adult lung in the basal layer of the upper airways, within or near pulmonary neuroendocrine cell rests, at the bronchoalveolar junction, as well as within the alveolar epithelium. These stem cells are hypothesized to be the source of lung regeneration and repair. But this mechanism seems to be insufficient after lung injury. There is increasing excitement over the last few years with the suggestion that exogenous stem cells may offer new treatment options for ARDS. Exogenous stem cells have the ability to differentiate and function as both airway and lung parenchymal epithelial cells in both in vitro and increasingly in vivo experiments. However, there is great controversy concerning the repair effect of adult stem cells in lung injury. This review evaluates the advances in endogenous respiratory stem cells, and assesses the evidence for the use of stem cells in the repair of lung injury.
Adult Stem Cells ; physiology ; transplantation ; Bone Marrow Transplantation ; Bronchi ; cytology ; Cell Fusion ; Epithelial Cells ; physiology ; Humans ; Pulmonary Alveoli ; cytology ; Respiratory Distress Syndrome, Adult ; therapy

Irreversible destruction of bronchi and alveoli can lead to multiple incurable lung diseases. Identifying lung stem/progenitor cells with regenerative capacity and utilizing them to reconstruct functional tissue is one of the biggest hopes to reverse the damage and cure such diseases. Here we showed that a rare population of SOX9 basal cells (BCs) located at airway epithelium rugae can regenerate adult human lung. Human SOX9 BCs can be readily isolated by bronchoscopic brushing and indefinitely expanded in feeder-free condition. Expanded human SOX9 BCs can give rise to alveolar and bronchiolar epithelium after being transplanted into injured mouse lung, with air-blood exchange system reconstructed and recipient's lung function improved. Manipulation of lung microenvironment with Pirfenidone to suppress TGF-β signaling could further boost the transplantation efficiency. Moreover, we conducted the first autologous SOX9 BCs transplantation clinical trial in two bronchiectasis patients. Lung tissue repair and pulmonary function enhancement was observed in patients 3-12 months after cell transplantation. Altogether our current work indicated that functional adult human lung structure can be reconstituted by orthotopic transplantation of tissue-specific stem/progenitor cells, which could be translated into a mature regenerative therapeutic strategy in near future.
Bronchiectasis ; genetics ; metabolism ; Humans ; Pulmonary Alveoli ; cytology ; metabolism ; SOX9 Transcription Factor ; genetics ; metabolism ; Stem Cell Transplantation ; methods ; Stem Cells ; cytology ; metabolism