Bleomycin-induced pulmonary fibrosis is independent of eosinophils

Eosinophils have been shown to increase in tissues during many fibrotic conditions and consequently have been suggested to contribute to the development of fibrosis. This study tested the hypothesis that eosinophils are essential in the development of lung fibrosis in mice in response to bleomycin (BLM). Anti-IL-5 antibody was administered intraperitoneally into mice 2 h prior to endotracheal BLM inoculation and thereafter, every other day. Lung eosinophilia was evaluated by measurement of eosinophil peroxidase activity and confirmed by eosinophil counts in histologic sections. Lung fibrosis was evaluated by hydroxyproline content and confirmed by collagen staining in histological sections. Results demonstrated that BLM induced pronounced lung eosinophilia, which was maximal 7 days after BLM treatment and remained elevated through day 14, in C57B1/6 SCID mice and CBA/J mice. In contrast, eosinophilia was a minor component in the lungs of wildtype C57B1/6 mice after BLM treatment, although lung fibrosis developed similarly in all three strains of mice. Treatment with anti-IL-5 completely abrogated eosinophilia but failed to block pulmonary fibrosis induced by BLM in all mouse strains, including C57B1/6 SCID, wildtype C57B1/6 mice, and CBA/J mice. Analysis of cytokine mRNA by RNase-protection assay in C57B1/6 SCID mice indicated that BLM treatment caused enhanced expression of the cytokines, TNF-alpha, and IL-6 at days 3, 7, and 14 post-BLM inoculation, regardless of whether eosinophils were depleted by anti-IL-5. Finally, the importance of eosinophils in lung fibrosis was examined in IL-5 gene knockout mice (IL-5tm1Kopf). BLM treatment induced significant lung fibrosis in IL-5 knockout mice in the absence of eosinophilia. These findings indicate that eosinophils are not an absolute requirement for BLM-induced pulmonary fibrosis in the mouse.     

Bleomycin-induced pulmonary fibrosis in genetically mast cell-deficient WBB6F1-W/Wv mice and mechanism of the suppressive effect of tranilast, an antiallergic drug inhibiting mediator release from mast cells, on fibrosis.

It has been well known that the number of mast cells increases during the development of fibrosis in various tissues including the lung. However, the role of mast cells in fibrosis still remains obscure. In the present paper, we evidenced that pulmonary fibrosis could be induced in genetically mast cell-deficient WBB6F1-W/Wv mice as well as WBB6F1-(+/+) mice having mast cells normally by the treatment with bleomycin (BLM, 5 mg/kg, i.v., 10 days), and there was not much difference in the histological changes of lungs between the two strains. An increase in the hydroxyproline content of the lung of WBB6F1-W/Wv mice was rather higher than that of WBB6F1-(+/+) mice. Previously, we reported that tranilast, an antiallergic drug inhibiting chemical mediator release from mast cells, suppressed the development of BLM-induced pulmonary fibrosis in ICR mice, suggesting the possibility that mast cells play certain roles in fibrosis. However, it was evidenced in the present report that tranilast suppressed BLM-induced fibrosis in WBB6F1-W/Wv mice. Tranilast neither suppressed the cytotoxic activity of BLM against KB cells and L-929 cells in vitro, nor inhibited the antitumor activity of BLM against Sarcoma-180 transplanted subcutaneously into ICR mice. Tranilast may act through suppressing BLM-induced activation of lymphoid cells including macrophage and neutrophil. These results indicate an inconsequential role of mast cells in the development of fibrosis. Increases in the number of mast cells and in histamine content of the lung, which were widely reported in the lungs of BLM-treated mice, may be the result of fibrosis.     

Metformin Reduces Bleomycin-induced Pulmonary Fibrosis in Mice

Metformin has anti-inflammatory and anti-fibrotic effects. We investigated whether metformin has an inhibitory effect on bleomycin (BLM)-induced pulmonary fibrosis in a murine model. A total of 62 mice were divided into 5 groups: control, metformin (100 mg/kg), BLM, and BLM with metformin (50 mg/kg or 100 mg/kg). Metformin was administered to the mice orally once a day from day 1. We sacrificed half of the mice on day 10 and collected the bronchoalveolar lavage fluid (BALF) from their left lungs. The remaining mice were sacrificed and analyzed on day 21. The right lungs were harvested for histological analyses. The messenger RNA (mRNA) levels of epithelial-mesenchymal transition markers were determined via analysis of the harvested lungs on day 21. The mice treated with BLM and metformin (50 mg/kg or 100 mg/kg) showed significantly lower levels of inflammatory cells in the BALF compared with the BLM-only mice on days 10 and 21. The histological examination revealed that the metformin treatment led to a greater reduction in inflammation than the treatment with BLM alone. The mRNA levels of collagen, collagen-1, procollagen, fibronectin, and transforming growth factor-β in the metformin-treated mice were lower than those in the BLM-only mice on day 21, although statistical significance was observed only in the case of procollagen due to the small number of live mice in the BLM-only group. Additionally, treatment with metformin reduced fibrosis to a greater extent than treatment with BLM alone. Metformin suppresses the inflammatory and fibrotic processes of BLM-induced pulmonary fibrosis in a murine model.     

Type II cell proliferation related to migration of inflammatory cells into the lung

Enhanced proliferation of epithelial cells in the lung is usually associated with some form of cellular injury. However, increased epithelial cell proliferation has also been observed in the absence of morphological signs of cellular injury. The nature of the stimuli to cell proliferation in the absence of cell injury is not clear. We hypothesized that one stimulus to epithelial cell proliferation in the absence of morphological injury to lung cells was the migration of inflammatory cells through the epithelium from the vasculature. This hypothesis was tested by comparing proliferation in the lungs of normal mice with that of white blood cell (WBC)-depleted mice when migration of WBC was stimulated by carbon instillation. White blood cell depletion in mice was accomplished with injections of 89SrCl2 (irradiated group). Sixteen days later, when WBC levels were reduced by about 64%, 4 mg colloidal carbon was instilled intratracheally into both normal and irradiated mice. At intervals of 1 through 4, 7, and 14 days after carbon instillation, four mice in each group were given ip tritiated thymidine and killed 1 hr later, and the lungs lavaged and prepared for electron microscopy and light microscope autoradiography. Analysis of lavage fluid revealed about 69% fewer cells in the irradiated mice compared to the normal mice at 1 day. There was no electron microscopic evidence of injury to the alveolar epithelium in either the normal or irradiated mice but there was increased proliferation of Type II cells. The proliferative response of Type II cells was 100 and 300% greater in the normal lung compared to irradiated mice at 3 and 4 days after carbon instillation, respectively. These data support the hypothesis that epithelial cell proliferation in the lung, in the absence of cellular injury, is related to the migration of WBC into the lung.     

Reactivation of Chlamydia trachomatis lung infection in mice by cortisone.

To study the latency, chronicity, and recurrent nature of chlamydial infection, we attempted to reactivate Chlamydia trachomatis lung infection in mice by immunosuppressive therapy with cortisone. Mice were treated with subcutaneous injections of cortisone acetate (125 mg/kg) every other day, starting on day 14 after intranasal inoculation of C. trachomatis serotype B (TW-5). C. trachomatis was recovered from the lungs beginning day 6 after the start of cortisone treatment until the end of the observation period on day 12 of treatment. Overall, the reactivation was successful in 8 of 55 mice treated with cortisone, in contrast to 0 of 41 inoculated, untreated mice (P = 0.009) and 0 of 35 uninoculated, treated mice. Cortisone treatment affected the ability of peritoneal exudate cells to respond to migratory inhibition after exposure to purified whole organisms of C. trachomatis serotype B (TW-5) but had little effect on serum antibody titers, indicating a possible role for cellular immunity in resistance against C. trachomatis infection in the lung.     

FTS reduces bleomycin-induced cytokine and chemokine production and inhibits pulmonary fibrosis in mice

Bleomycin (BLM), an antitumour drug, is known to cause interstitial pneumonia followed by pulmonary fibrosis, and has often been used to produce an animal model of pulmonary fibrosis. In the present study, we examined the effect of a nonapeptide thymic hormone, facteur thymique serique (FTS), on the murine lung fibrosis induced by intratracheal instillation of BLM. Treatment with FTS ameliorated BLM-induced fibrotic changes in a dose-dependent manner, as indicated by the reduced accumulation of hydroxyproline (HP). In addition, FTS suppressed BLM-induced cellular inflammatory response in the lungs, as evidenced by inhibition of increased lung weight, reduced accumulation of inflammatory leucocytes, including lymphocytes and neutrophils, but not macrophages, and less pronounced histopathological changes. Finally, BLM challenge increased the local synthesis of proinflammatory cytokines, TNF-alpha and IL-1beta and chemokines, MCP-1, MIP-1alpha RANTES, MIP-2 and KC, while administration of FTS suppressed the production of these cytokines, except for MCP-1. These effects of FTS were observed only when mice received intratracheal instillation with BLM. Considered collectively, our results indicated that FTS treatment ameliorated the cellular inflammatory responses and fibrotic changes in the lungs caused by BLM and such inhibition was well correlated with reduced synthesis of several fibrosis-related cytokines, and suggested that FTS may be potentially useful for the treatment of pulmonary fibrosis.     

Effects of granulocyte colony-stimulating factor on the kinetics of inflammatory cells in the peripheral blood and pulmonary lesions during the development of bleomycin-induced lung injury in rats

We evaluated the effects of granulocyte colony-stimulating factor (G-CSF) on the kinetics of inflammatory cells during the development of inflammation in bleomycin (BLM)-induced lung injury. G-CSF (100 microg/kg/day, s.c.) was administered to rats treated with or without BLM (2 mg/200 microl, intratracheally) for up to 14 days (Day 14) immediately after BLM treatment. In the BLM + G-CSF group, the lung injury score increased on Days 1 and 14, and the score of lung fibrosis on Day 14, respectively. Except for neutrophils, there were no effects of G-CSF on the number of inflammatory cells both in the peripheral blood and in the lung in both BLM-treated and -untreated rats at the acute inflammatory phase. In the G-CSF-treated groups, the number of neutrophil counts in the peripheral blood drastically increased on Day 1, temporally decreased on Day 3, and increased again on Days 7 and 14. The number of neutrophils in the lung markedly increased on Day 1 and then remained at a plateau level until Day 14. The neutrophil alkaline phosphatase score in the lung commenced to increase on Day 1, reached the maximal level on Day 7, and then remained at a plateau level until Day 14. Correlations between the numbers of neutrophils in the lung and the peripheral blood or the lung lesion score were only observed on Day 14. These findings suggest that the exacerbating effect of G-CSF on the lung injury coincided with the increase in the number of alkaline phosphatase-positive neutrophils infiltrating in the pulmonary lesion at the acute inflammatory phase and it lasted to the fibrogenic phase. The exacerbating effect of G-CSF on the severe BLM-induced lung injury seems to be related not only to the pulmonary accumulation of activated neutrophils but also to the severity of lung injury caused by the direct effects of BLM.     

Response of lung gammadelta T cells to experimental sepsis in mice

Gammadelta T cells link innate and adaptive immune systems and may regulate host defence. Their role in systemic inflammation induced by trauma or infection (sepsis) is still obscured. The present study was aimed to investigate functions of lung gammadelta T cells and their response to experimental sepsis. Mice were subjected to caecal ligation and puncture (CLP) to induce sepsis and acute lung injury (ALI), or to the sham operation. Animals were killed 1, 4, and 7 days postoperatively; lungs were examined by histology, and isolated cells were studied by flow cytometry. Absolute number of gammadelta T cells progressively increased in lungs during sepsis, and reached a seven-fold increase at day 7 after CLP (3.84 +/- 0.41 x 10(5)/lung; P = 0.0002 versus sham). A cellular dysfunction was revealed one day after CLP, as manifested by low cytolytic activity (22.3 +/- 7.1%; P < 0.05 versus sham), low interferon-gamma (IFN-gamma; 8.5 +/- 2.5%; P < 0.05 versus control) and interleukin-10 (IL-10) expression, and high tumour necrosis factor-alpha expression (19.5 +/- 1.7%; P < 0.05 versus control). The restoration of cytotoxicity, and increase in IFN-gamma and IL-10 expression was observed at day 7 of CLP-induced sepsis. In summary, our results demonstrate significant progressive accumulation of gammadelta T cells in lungs during CLP-induced ALI. The temporary functional suppression of lung gammadelta T cells found early after CLP may influence the outcome of sepsis, possibly being associated with uncontrolled inflammatory lung damage.     

Poly(ADP-ribose) synthetase activity during bleomycin-induced lung fibrosis in hamsters

Bleomycin damages cellular DNA and is a potent inducer of pulmonary fibrosis. It has been shown to act through a superoxide-mediated mechanism. We are interested in determining the biochemical mechanisms involved in fibrosis and in this preliminary study we have examined the temporal relationship between early biochemical events associated with DNA damage and fibrosis, in lungs of hamsters after administration of 0.75 unit of bleomycin. The activities of poly(ADP-ribose) synthetase, an enzyme associated with DNA repair, inducible superoxide dismutase (SOD) and prolyl hydroxylase as well as the tissue levels of NAD+ and hydroxyproline in the lung were determined. All three enzyme activities expressed as per milligram DNA or per lung, increased upon bleomycin treatment over the saline-administered controls. Lung poly(ADP-ribose) synthetase activity which is sensitive to DNA breaks, increased first (24% over control in 1 day, P less than 0.0001), attained the maximum value on the 5th day (952% over control, P less than 0.0001), and started to decline thereafter and approached near the control value on 14th day. Bleomycin treatment induced a rapid change in the level of lung NAD+. After 1 day the level of NAD+ was reduced by 42% compared to the control (P less than 0.001), further declined to 65% (P less than 0.001) on the 3rd day, and stayed at that level until the 7th day. On the 14th day, however, the NAD+ level was still lower (29%, P less than 0.05) but approaching the value in the control animals. The activity of prolyl hydroxylase showed significant increase on the 3rd day (50% over control, P less than 0.0001) after bleomycin administration. The enzyme activity continued to increase until the end of the experiment (490% of control, P less than 0.0001, on Day 14). The content of undialyzable hydroxyproline, a marker for collagen, was also increased significantly in the lung tissue on the 3rd day (30% over control, P less than 0.05), continued to increase and reached the highest level on the 14th day (71% over control, P less than 0.001). A significant increase in the activity of SOD (19% over control, P less than 0.001) was seen on the 5th day which continued to increase and attained the highest value on Day 14 (115% over control, P less than 0.0001).(ABSTRACT TRUNCATED AT 400 WORDS)     

Activation of Valpha14(+) natural killer T cells by alpha-galactosylceramide results in development of Th1 response and local host resistance in mice infected with Cryptococcus neoformans

We examined the effect of alpha-galactosylceramide (alpha-GalCer) on the synthesis of gamma interferon (IFN-gamma) and local resistance in mice infected intravenously with Cryptococcus neoformans. The level of IFN-gamma in serum increased on day 3, reached a peak level on day 7, and decreased to the basal level on day 14 postinfection in mice treated with alpha-GalCer, while in vehicle-treated mice, no increase was detected at any time points except for a small increase on day 7. Such effects were not observed in NKT-KO mice. In CD4KO mice, minor synthesis of IFN-gamma was detected on day 3 in sera but was completely abolished by day 7. The alpha-GalCer-induced IFN-gamma production on day 3 was partially reduced in mice depleted of NK cells by treatment with anti-asialo-GM(1) antibody (Ab). Spleen cells obtained from infected and alpha-GalCer-treated mice on day 7 produced a large amount of IFN-gamma upon restimulation with live organisms, while only a marginal level of production was detected in splenocytes from infected and vehicle-treated mice. Such effects were abolished in CD4KO and NKT-KO mice. Finally, the fungal loads in the lungs and spleen on days 7 and 14 were significantly reduced in alpha-GalCer-treated mice compared to those in control mice. In NKT-KO mice, local resistance elicited by alpha-GalCer was completely abolished, although no obvious exacerbation of infection was detected. Furthermore, treatment with anti-IFN-gamma monoclonal Ab mostly abrogated the protective effect of this agent. Thus, our results indicated that activation of Valpha14(+) NKT cells resulted in an increased Th1 response and local resistance to C. neoformans through production of IFN-gamma.     

Effects of pretreatment with androgen or thyrold hormone on androgen-induced proliferation of granular convoluted tubular cells in mouse submandibular glands

The effects of pretreatment with androgen or thyroid hormone on androgen-induced proliferation of granular convoluted tubular cells (GCT cells) in the submandibular glands of ovariectomized female BALB/c or C57BL/6 mice were investigated. The proliferation of GCT cells was estimated by their labeling index. Daily injections of 5α-dihydrotestosterone (DHT) (100 μg/mouse/day) caused a transient increase in the labeling index of GCT cells of ovariectomized 60-day-old BALB/c mice during the first four injections, but injections of thyroxine (T₄) (15 μg/mouse/day) did not. On the other hand, both DHT and T₄ increased the esteroprotease activity, a marker of the differentiation of GCT cells, time dependently. Injections of DHT into ovariectomized 102-day-old BALB/c mice also caused a transient increase in the labeling index of GCT cells. However, pretreatment of ovariectomized 60-day-old BALB/c mice with DHT for 4 or 14 days completely abolished the DHT-induced increase in the labeling index of 102-day-old mice, and pretreatment with T₄ for 14 days reduced this increase. Pretreatment with DHT or T₄ for 14 days did not affect the DHT-induced increase in esteroprotease activity. Pretreatment of ovariectomized 60-day-old C57BL/6 mice with DHT for 14 days also completely abolished the DHT-induced increase in the labeling index of GCT cells at the age of 102 days, but pretreatment with T₄ for 14 days did not affect the increase. These results suggest that the proliferation of mouse GCT cells induced by androgen results in a complete abolition of their proliferative response to androgen, and that the effect of thyroid hormone on the proliferative response of GCT cells to androgen may differ in different strains of mice. © 1993 Wiley-Liss, Inc.     

The acute effects of single and repeated injections of acrolein and other aldehydes

The irritating aldehyde acrolein was injected intraperitoneally into mice. A single injection at 4 mg/kg gave rise to a 5-fold increase in plasma total lactate dehydrogenase (LDH) activity, with the peak after approximately 10 h. The pattern of LDH isoenzymes was not altered. Repeated injections (daily or weekly) caused a progressively less pronounced effect on the LDH activity. Experiments with formaldehyde and crotonaldehyde gave essentially the same results. The LD50 for acrolein i.p. in mice was increased from a level of 7 mg/kg to a level of 12 mg/kg by pretreatment with sublethal doses of 4 mg/kg/day for 5 days. Thus, the response to repeated acrolein injections, in terms of LDH and LD50, indicates an acquired tolerance against the irritant. Likewise, pretreatment with formaldehyde or crotonaldehyde could induce tolerance, in terms of LDH activity, towards a subsequent injection of acrolein. Histopathological examination revealed that spleen, adrenals and thymus were affected. The thymus markedly decreased in size after repeated injections of acrolein, crotonaldehyde or formaldehyde. Adrenalectomized mice given acrolein showed no thymus atrophy. A single injection of aldehyde caused an increased level of the adrenal hormone corticosterone in blood plasma. Adrenalectomized mice still showed a certain tolerance, in terms of LDH activity, after repeated injections of acrolein, but the increase in plasma LDH activity was smaller than for normal animals. Treatment with acrolein for six days did not change the level of reduced glutathione or the glutathione S-transferase activity in liver cytosol, but the rate of glutathione synthesis was increased. It is concluded that adrenalectomy does not completely prevent the development of tolerance in mice. It is possible that an increased metabolism can partially explain the acquired tolerance.     

Cellular changes in the lungs of adrenalectomized rats following left pneumonectomy

The time course and nature of the cellular response to left pneumonectomy, with or without prior adrenalectomy, were evaluated in the right lungs of male Sprague-Dawley rats using morphometric techniques. Animals were studied at days 2, 5, and 14 following pneumonectomy, intervals prior to, during the course of, and following significant compensatory changes in right lung mass. The postoperative increase in right lung mass and volume in pneumonectomized animals involved minimal changes in the ratios of most tissue components, when compared to the lungs of sham-operated controls. A transient disproportionate increase in type II cell volume and epithelial thickness was evident on day 14. Postpneumonectomy changes in the type II epithelium were accentuated in the lungs of adrenalectomized-pneumonectomized animals. Adrenalectomy 5 days prior to pneumonectomy resulted in a substantial increase in the volume of all right lung tissue components, associated with thickening of the alveolar wall and with increases in the volume of both cellular and noncellular interstitium. Effects of adrenalectomy on the endothelium also were evident. In both adrenal-intact and adrenalectomized animals, pneumonectomy increased alveolar number by day 14 but had no effect on the volume of individual alveoli. These results confirm a coordinated pattern of compensatory growth following pneumonectomy in the adrenal-intact rat. The data further suggest that in adrenalectomized animals compensatory lung growth is more poorly synchronized, with pronounced postoperative elevations in volume of the interstitial and type II epithelial compartments leading to increased thickness of the alveolar wall. Adrenal hormones thus appear to be required for coordination and control of compensatory lung growth and for rapid restoration of normal tissue structure.     

Neutralization of transforming growth factor-beta 1 in a mouse model of immune-induced lung fibrosis.

We examined the contribution of the cytokine transforming growth factor beta 1 (TGF-beta 1) in the inflammatory response and fibrotic reaction in a mouse model of immune-induced lung fibrosis caused by repeated intranasal exposure to heat-killed bacillus Calmette-Guérin (BCG). Mice received 200 micrograms of BCG 3 days/week for 4 weeks, and simultaneous intraperitoneal injections of a monospecific rabbit antiserum against mouse TGF-beta 1 or a preimmune serum (normal rabbit globulin). BCG instillations generated a copious release of antigenic TGF-beta 1 in the lungs at 1, 2, 3 and 4 weeks (up to 15 ng/lungs/mouse). Treatment with anti-TGF-beta 1 antiserum significantly diminished the number of free lung cells recovered by bronchoalveolar lavage (BAL), although the BAL cellular profile was not affected. Moreover, anti-TGF-beta 1 treatment of challenged mice diminished very significantly the total levels of interleukin-1 beta (IL-1 beta) and tumour necrosis factor-alpha (TNF-alpha) in the lungs of animals challenged with BCG. Histological examination and morphometric analysis of Masson's Trichrome-stained sections and measurements of total lung hydroxyproline levels showed a substantial decrease in lung fibrosis and granulomatous response of challenged mice given anti-TGF-beta 1. These data argue for a role for TGF-beta 1 in inducing inflammation and lung fibrosis in response to an immune stimulus.     

Treatment with inhaled formulation of angiotensin-(1-7) reverses inflammation and pulmonary remodeling in a model of chronic asthma

Asthma is characterized by inflammation, pulmonary remodeling and bronchial hyperresponsiveness. We have previously shown that treatment with angiotensin-(1-7) [Ang-(1-7)] promotes resolution of eosinophilic inflammation and prevents chronic allergic lung inflammation. Here, we evaluated the effect of treatment with the inclusion compound of Ang-(1-7) in hydroxypropyl β-cyclodextrin (HPβCD) given by inhalation on pulmonary remodeling in an ovalbumin (OVA)-induced chronic allergic lung inflammation. Mice were sensitized to ovalbumin (OVA; 4 injections over 42 days, 14 days apart) and were challenged 3 times per week, for 4 weeks (days 21–46). After the 2nd week of challenge, mice were treated with Ang-(1-7) by inhalation (4.5 μg of Ang-(1-7) included in 6.9 μg of HPβCD for 14 days, i.e. days 35–48). Mice were killed 72 h after the last challenge and blood, bronchoalveolar lavage fluid (BALF) and lungs were collected. Histology and morphometric analysis were performed in the lung. Metalloproteinase (MMP)-9 and MMP-12 expression and activity, IL-5, CCL11 in the lung and plasma IgE were measured. After 2 weeks of OVA challenge there was an increase in plasma IgE and in inflammatory cells infiltration in the lung of asthmatic mice. Treatment with inhaled administration of Ang-(1-7)/HPβCD for 14 days reduced eosinophils, IL5, CCL11 in the lung and plasma IgE. Treatment of asthmatic mice with Ang-(1-7)/HPβCD by inhalation reversed pulmonary remodeling by reducing collagen deposition and MMP-9 and MMP-12 expression and activity. These results show for the first time that treatment by inhalation with Ang-(1-7) can reverse an installed asthma, inhibiting pulmonary inflammation and remodeling.     

小鼠实验性肺纤维化上皮细胞-间充质细胞转变及骨桥蛋白的表达

目的从形态学上证实小鼠实验性肺纤维化上皮细胞-间充质细胞转变(EMT)及骨桥蛋白(OPN)的表达变化。方法将60只健康雄性C57BL/6小鼠随机分为生理盐水对照组和博来霉素(BLM)模型组,采用口咽抽吸法经咽部注入气管50μl博来霉素溶液,对照组注入50μl生理盐水。分别在造模后第3d、7d、14d、21d及28d 5个时间点处死对照组及模型组小鼠。取小鼠左肺制备石蜡切片,行苏木素-伊红(HE)染色及天狼猩红染色观察肺组织形态变化及胶原增生情况;行表面活性物质相关蛋白-C(SP-C)、上皮型钙黏蛋白(E-Cadherin)、波形蛋白(vimentin)、成纤维细胞特异蛋白1(FSP1)和骨桥蛋白(OPN)免疫组织化学染色,观察肺纤维化上皮细胞-间充质细胞转变(EMT)形态与OPN表达,利用图像分析软件Image-Pro Plus 6.0(IPP6.0)测量切片积分吸光度(IA),并进行统计分析。结果HE及天狼猩红染色显示出小鼠肺组织正常结构和纤维化形态变化,并且模型组小鼠肺组织的胶原生成量随时间推移而增加(P<0.05)。免疫组织化学结果显示,模型组小鼠肺OPN表达量随病程进展持续升高,在纤维化期达到峰值,而对照组表达量很少(P<0.05);对照组SP-C在Ⅱ型肺泡细胞恒定表达,模型组SP-C持续增加,且胞体增大;对照组E-Cadherin表达量多,而模型组在肺实变部位几乎不表达,但肺实变部位产生波形蛋白及FSP1,这些蛋白提示肺纤维化存在EMT。结论BLM诱导性肺纤维化中存在EMT,且与显著增加的OPN有潜在关系。     

Correction of L-NAME-induced disturbances in DNA synthesis and free radical oxidation in respiratory organs of newborn albino rats with dalargin

Intraperitoneal injections of L-NAME (5 injections, 9.3×10^-5 mol/kg each) to albino rats from the 2nd to 6th day of life inhibited DNA synthesis in epithelial cells, stimulated this process in bronchial smooth muscle cells, and intensified free radical oxidation in the lungs. Dalargin administered in a dose of 1.4×10^-7 mol/kg 30 min after treatment with NG-nitro-L-arginine methyl ester abolished its effect on epithelial cells and attenuated changes in smooth muscle cells. Correcting activity of dalargin is associated with its antiradical and antioxidant properties.     

Dynamics of the content and synthesis of nucleic acids and proteins in the lungs during adaptation to hypoxia

The relative weight of the lungs and the synthesis of nucleic acids and protein in the lungs of rats adapted to hypoxia were studied. Adaptation was carried out for 6 h a day in a pressure chamber at an “altitude” of 7000 m for 40 days. An increase in the relative weight of the lungs, in the relative (per 100 mg lung tissue) and total RNA content, and in the intensity of protein synthesis was found. The relative DNA content (per 100 mg) in the lung tissue was unchanged.     

Adaptive responses of the pulmonary macrophagic system to carbon. I. Kinetic studies.

The adaptive capacity of the lung to increase the output of alveolar macrophages is vital to its handling of foreign material, particularly in an overload situation. In previous papers we demonstrated the role of pulmonary intersititial cells in providing a pool of precursor cells that may be available for this purpose. We now extend these observations to a study of pulmonary cytodynamics in mice subjected to a single large load (4 mg.) of carbon delivered by endotracheal tube. The output of free macrophages over a period of 14 days was correlated with DNA synthesis of pulmonary cells as measured by their uptake of 3H-thymidine. The initial increase of macrophagic output, 5 times in 12 hours and 10 times in 24 hours occurred before any increase in mitotic activity or cellularity was demonstrated in the pulmonary interstitium. The high level of macrophagic cell output which was maintained over the next week was accompanied by increased thymidine uptake in the lung. Increased mitotic activity was confined to the interstitial cell population; no mitoses were observed in free macrophages. The results indicate a biphasic macrophagic response to inhaled particulates, an early phase apparently unrelated to a local cellular response and a later phase of interstitial cell proliferation which appears to be concerned with the maintenance of the high output of macrophages.