Modulation of ovalbumin-induced airway inflammation and hyperreactivity by tolerogenic APC

Allergic asthma is mediated in part by unregulated Th2 inflammation in response to an allergen. Induction of peripheral tolerance by inoculation of Ags into the anterior chamber of the eye (ocular tolerance) before sensitization blocks Th2 responses. Thus, we proposed that induction of ocular tolerance to the allergen might modulate an ongoing allergen-induced Th2 pathogenesis in the lung. We initiated ocular tolerance in previously immunized mice in a classic mouse model of OVA-induced pulmonary allergic inflammation. In the model of ocular tolerance, the need for inoculation of Ag into the anterior chamber can be bypassed by i.v. inoculation of in vitro-generated tolerogenic (TGF-beta2-treated, Ag-pulsed) APC (tol-APC). We observed that with i.v. inoculation, such tolerogenic APC, but not control APC, reduced eosinophil and lymphocyte pulmonary infiltration in experimental mice. Similarly, production of Th2 cytokines (IL-4, -5, and -13), but not IFN-gamma, was reduced. Importantly, airway hyperresponsiveness and mucus production were significantly reduced after treatment with the tol-APC. We also show that in vitro suppression of IL-13 production from OVA-sensitized effector T cells was mediated by CD8+, not CD4+, T regulatory cells. Thus, i.v. inoculation of the tol-APC induced peripheral tolerance that suppressed Th2-mediated pathogenesis in the lungs of presensitized mice. The ability of the tol-APC to induce peripheral tolerance and suppress existing Th2 immune inflammation may lead to novel therapies for pulmonary allergic inflammation and its related pathology.     

CD40/CD40 ligand interactions are critical for elicitation of autoimmune-mediated fibrosis in the lung

Pulmonary interstitial fibrosis (PIF), associated with persistent inflammation and increased collagen deposition in the interstitium, is often considered an autoimmune disease. Hapten immune PIF (HIPIF), a model for PIF, is elicited in the lung by a single intratracheal (i.t.) challenge in mice sensitized with hapten (2,4,6-trinitrobenzene sulfonic acid, TNBS). In this study, we characterized the role of CD40/CD40 ligand (CD40L) interactions in the elicitation of secondary cell-mediated immune responses that lead to development of fibrosis in the lung using an adoptive transfer model of HIPIF. The expression of CD40 was detected on bronchoalveolar lavage (BAL) cells 1-3 days after i.t. challenge with hapten in the HIPIF lung, but not lungs from the control mice. The CD40(bright) BAL cells morphologically resembled infiltrating monocytes. Furthermore, blocking CD40/CD40L interactions with blocking Ab decreased BAL production of Th1-mediators (IL-12 and TNF-alpha). Moreover, either blocking CD40/CD40L interactions with the Ab or using IL-12 knockout recipient mice prevented the increased collagen deposition (accumulation of hydroxyproline) in the lungs during HIPIF induction. We conclude that second signals (CD40/CD40L interactions) are required for elicitation of secondary immune responses that lead to PIF in vivo. The results support the notion that CD40/CD40L interactions are involved in the pathogenesis of an ongoing autoimmune disease.     

Immunological tolerance to hapten prevents subsequent induction of hapten-immune pulmonary interstitial fibrosis (HIPIF).

Pulmonary interstitial fibrosis (PIF) is a morphological term which in part can be defined as accumulation of collagen in the extracellular matrix. Previously we showed that hamsters sensitized with 2,4,6-trinitro-1-chlorobenzene (TNCB) developed PIF 14 days after an intratracheal challenge with 2,4,6-trinitrobenzene sulfonic acid (TNBS). The participation of delayed-type hypersensitivity (DTH) in lung collagen deposition was clearly demonstrated. In this paper, we use an adaptation of this model to mice and show that the lung collagen deposition observed was related to the genetic ability of the strain to maintain a DTH response to the immunizing hapten (TNP). Specifically, the lung collagen deposition on Day 14 in hapten-sensitized, challenged animals in high responder to TNP (BALB/c, H-2d) was higher than that in low responder mouse (C57BL/6, H-2b). Furthermore, aged C57BL/6 strain (retired breeders) possessed a DTH response to TNP and produced significantly higher accumulation of hydroxyproline than that of TNBS-challenged-only animals. A DTH mechanism for the induction of the fibrosis is consistent with the observation that responder mice that were made tolerant to the antigen were unable to respond to the lung challenge with a specific increase in lung index or collagen deposition. These results suggest that effector T lymphocytes that are important in DTH play a key role in the regulation of lung collagen deposition in hapten-immune pulmonary interstitial fibrosis (HIPIF) in mice.     

Allergic contact dermatitis induced by intratracheal administration of hapten

It is known that the skin is the natural route for induction of allergic contact dermatitis (ACD). Because the lung is another potential portal of entry for sensitizing chemicals, studies were performed to evaluate immune responses to haptens deposited in the lung. As a result, a new method for inducing ACD was developed. Intratracheal (IT) inoculation of 2,4,6-trinitrobenzene sulfonic acid (TNBS) led to maximal ear swelling 24 hr after challenge on the ear with 2,4,6-trinitrochlorobenzene (TNCB) in carrier. This response was specific for immunizing hapten. Furthermore, it was equally possible to induce ACD by intratracheal inoculation of trinitrophenyl- (TNP) modified bronchoalveolar cells (BAC) or haptenated spleen cells. Adoptive transfer studies demonstrated that the hypersensitivity that resulted from IT inoculation of TNBS or TNP BAC could be transferred with cells. With a monoclonal anti-hamster Ia reagent, it was demonstrated that, like Langerhans cells, 80 to 90% of cells in the bronchial lavage fluid expressed Ia determinants on their membrane. These cells were morphologically indistinguishable from macrophages. Because an individual is capable of being sensitized when hapten is introduced by the pulmonary or epicutaneous routes, the possibility is raised that the alveolar macrophages in the lung possess a similar capability for antigen presentation of hapten in the induction of ACD as does the Langerhans cell.     

Induction of IL-13 triggers TGF-beta1-dependent tissue fibrosis in chronic 2,4,6-trinitrobenzene sulfonic acid colitis

To investigate the immunopathogenesis of inflammation-associated fibrosis, we analyzed the chronic colitis and late-developing fibrosis occurring in BALB/c mice administered weekly doses of intrarectal 2,4,6-trinitrobenzene sulfonic acid. We showed first in this model that an initial Th1 response involving IL-12p70 and IFN-gamma subsides after 3 wk to be supplanted by an IL-23/IL-25 response beginning after 4-5 wk. This evolution is followed by gradually increasing production of IL-17 and cytokines ordinarily seen in a Th2 response, particularly IL-13, which reaches a plateau at 8-9 wk. In vitro stimulation studies suggest that this IL-13 production is dependent on IL-23 and IL-25, but not on IL-12p70. We then show that IL-13 production results in the induction of an IL-13R formerly thought to function only as a decoy receptor, IL-13Ralpha(2), and this receptor is critical to the production of TGF-beta(1) and the onset of fibrosis. Thus, if IL-13 signaling through this receptor is blocked by administration of soluble IL-13Ralpha(2)-Fc, or by administration of IL-13Ralpha(2)-specific small interfering RNA, TGF-beta(1) is not produced and fibrosis does not occur. These studies show that in chronic 2,4,6-trinitrobenzene sulfonic acid colitis, fibrosis is dependent on the development of an IL-13 response that acts through a novel cell surface-expressed IL-13R to induce TGF-beta(1). A similar mechanism may obtain in certain forms of human inflammatory bowel disease.     

CD4+ NKT cells,but not conventional CD4+ T cells,are required to generate efferent CD8+ T regulatory cells following antigen inoculation in an immune-privileged site

Following inoculation of Ag into the anterior chamber (a.c.), systemic tolerance develops that is mediated in part by Ag-specific efferent CD8(+) T regulatory (Tr) cells. This model of tolerance is called a.c.-associated immune deviation. The generation of the efferent CD8(+) Tr cell in a.c.-associated immune deviation is dependent on IL-10-producing, CD1d-restricted, invariant Valpha14(+) NKT (iNKT) cells. The iNKT cell subpopulations are either CD4(+) or CD4(-)CD8(-) double negative. This report identifies the subpopulation of iNKT cells that is important for induction of the efferent Tr cell. Because MHC class II(-/-) (class II(-/-)) mice generate efferent Tr cells following a.c. inoculation, we conclude that conventional CD4(+) T cells are not needed for the development of efferent CD8(+) T cells. Furthermore, Ab depletion of CD4(+) cells in both wild-type mice (remove both conventional and CD4(+) NKT cells) and class II(-/-) mice (remove CD4(+) NKT cells) abrogated the generation of Tr cells. We conclude that CD4(+) NKT cells, but not the class II molecule or conventional CD4(+) T cells, are required for generation of efferent CD8(+) Tr cells following Ag introduction into the eye. Understanding the mechanisms that lead to the generation of efferent CD8(+) Tr cells may lead to novel immunotherapy for immune inflammatory diseases.     

Opposing regulatory roles of complement factor 5 in the development of bleomycin-induced pulmonary fibrosis

The mechanisms of idiopathic pulmonary fibrosis pathogenesis, a chronic and progressive interstitial lung disease, remain elusive. The complement system, a crucial arm of the innate immune response, plays a pivotal role in several pathological disorders; however, the contribution of individual complement components to lung fibrosis has not yet been examined. Complement factor 5 (C5) and its cleavage product C5a are critical mediators in inflammatory diseases. Thus, to evaluate the role of C5 in lung fibrosis, we compared congenic C5-sufficient and C5-deficient mice in a well-characterized murine model of bleomycin-induced pulmonary fibrosis. C5-deficient mice had an exaggerated inflammatory phenotype compared with C5-sufficient mice during acute bleomycin-induced lung injury. These findings suggest a protective and anti-inflammatory role for C5, which was linked to the regulation of matrix metalloproteinases involved in cell migration. In contrast, C5 had a detrimental effect during chronic stages of bleomycin-induced injury, indicating a profibrotic role for C5. This deleterious activity for C5 was associated with expression of the fibrogenic cytokine TGF-beta1 and matrix metalloproteinase-3, an important mediator in fibroblast contraction. Altogether, our data reveal novel and opposing roles for C5 in both inflammation and tissue repair. Furthermore, these findings provide insight into the development of new therapeutic strategies for idiopathic pulmonary fibrosis patients.     

Eosinophils and T lymphocytes possess distinct roles in bleomycin-induced lung injury and fibrosis

Leukocyte infiltration is characteristic of lung injury and fibrosis, and its role during tissue repair and fibrosis is incompletely understood. We found that overexpression of IL-5 in transgenic mice (IL-5(TG)) or by adenoviral gene transfer increased bleomycin (blm)-induced lung injury, fibrosis, and eosinophilia. Surprisingly, blm-treated IL-5-deficient (IL-5(-/-)) mice also developed pronounced pulmonary fibrosis but characterized by marked T lymphocyte infiltration and absence of eosinophilia. In both murine strains however, induction of lung TGF-beta expression was evident. Purified lung eosinophils from blm-treated IL-5(TG) mice stimulated alpha-smooth muscle actin and collagen expression in mouse lung fibroblasts, without affecting proliferation. Furthermore instillation of purified eosinophils into murine lungs resulted in extension of blm-induced lung fibrosis, thus confirming a role for eosinophils. However, lung T lymphocytes from blm-treated IL-5(-/-) mice were able to stimulate fibroblast proliferation but not alpha-smooth muscle actin or collagen expression. Blocking T cell influx by anti-CD3 Abs abrogated lung fibrosis, thus also implicating T lymphocytes as a key participant in fibrosis. Pulmonary fibrosis in IL-5(TG) mice was preferentially associated with type 2 cytokines (IL-4 and IL-13), whereas fibrotic lesions in IL-5(-/-) animals were accompanied by proinflammatory cytokine (TNF-alpha, IL-1beta, and IFN-gamma) expression. We suggest that eosinophils and T cells contribute distinctly to the development of blm-induced lung fibrosis potentially via their production of different cytokine components, which ultimately induce TGF-beta expression that is intimately involved with the fibrosis.     

TGF-beta 1 as an enhancer of Fas-mediated apoptosis of lung epithelial cells

Transforming growth factor-beta 1 (TGF-beta 1) has important roles in lung fibrosis and the potential to induce apoptosis in several types of cells. We previously demonstrated that apoptosis of lung epithelial cells induced by Fas ligation may be involved in the development of pulmonary fibrosis. In this study, we show that TGF-beta1 induces apoptosis of primary cultured bronchiolar epithelial cells via caspase-3 activation and down-regulation of cyclin-dependent kinase inhibitor p21. Concentrations of TGF-beta 1 that were not sufficient to induce apoptosis alone could enhance agonistic anti-Fas Ab or rFas ligand-mediated apoptosis of cultured bronchiolar epithelial cells. Soluble Fas ligand in the bronchoalveolar lavage fluid (BALF) from patients with idiopathic pulmonary fibrosis (IPF) also induced apoptosis of cultured bronchiolar epithelial cells that was significantly attenuated by anti-TGF-beta Ab. Otherwise, BALF from patients with hypersensitivity pneumonitis (HP) could not induce apoptosis on bronchiolar epithelial cells, despite its comparable amounts of soluble Fas ligand. The concentrations of TGF-beta 1 in BALF from patients with IPF were significantly higher compared with those in BALF from patients with HP or controls. Furthermore, coincubation with the low concentration of TGF-beta 1 and HP BALF created proapoptotic effects comparable with the IPF BALF. In vivo, the administration of TGF-beta 1 could enhance Fas-mediated epithelial cell apoptosis and lung injury via caspase-3 activation in mice. Our results demonstrate a novel role of TGF-beta 1 in the pathophysiology of pulmonary fibrosis as an enhancer of Fas-mediated apoptosis of lung epithelial cells.     

T cell-derived IL-10 promotes lung cancer growth by suppressing both T cell and APC function.

We have found previously that human lung cancers potently induce T lymphocyte IL-10 production in vitro. To assess the impact of enhanced T cell-derived IL-10 on antitumor immunity in vivo, we utilized transgenic mice expressing IL-10 under the control of the IL-2 promoter. We have shown previously that Lewis lung carcinoma cells (3LL) have more aggressive growth potential in IL-10 transgenic mice compared with control littermates. In this study, we show that transfer of T cells from IL-10 transgenic mice to control littermates transferred the IL-10 immunosuppressive effect and led to enhanced 3LL tumor growth. In addition to changes in T cell-mediated immunity, professional APC from IL-10 transgenic mice were found to have significantly suppressed capacity to induce MHC alloreactivity, CTL responses, and IL-12 production. Tumor Ag-pulsed dendritic cells from IL-10 transgenic mice also failed to generate antitumor reactivity. These results suggest that increased levels of T cell-derived IL-10 severely impair antitumor immunity in vivo, due to defects in both T cell and APC function.     

Reduction of Smad3 accelerates re-epithelialization in a murine model of colitis

To determine the role of Smad3 in re-epithelialization and inflammation, experimental colitis was induced in Smad3 heterozygous mice and their wild-type littermates by single intrarectal administration of 2,4,6-trinitrobenzene sulfonic acid (TNBS) in ethanol. The area of epithelial deficiency was significantly reduced in the heterozygotes on the 4th-6th day after TNBS administration as compared to the controls although the number of inflammatory cells in the colonic mucosa in the heterozygotes and their wild-type littermates varied similarly throughout the course of colitis. Proliferation of the intestinal epithelium in the heterozygotes was significantly accelerated as compared to that in the wild-type controls on the 1st and 2nd days after TNBS administration. These results suggest that reduction of Smad3 significantly accelerates re-epithelialization of the intestinal mucosa without enhancing inflammation. Suppression of TGF-beta1 induction in the colonic mucosa of the heterozygotes may lead to a higher level of proliferation of intestinal epithelial cells.     

Reversal of CD8+ T cell ignorance and induction of anti-tumor immunity by peptide-pulsed APC

In the present report, we have studied the potential of naive and activated effector CD8(+) T cells to function as anti-tumor T cells to a solid tumor using OVA-specific T cells from TCR-transgenic OT-I mice. Adoptive transfer of naive OT-I T cells into tumor-bearing syngeneic mice did not inhibit tumor cell growth. The adoptively transferred OT-I T cells did not proliferate in lymphoid tissue of tumor-bearing mice and were not anergized by the tumor. In contrast, adoptive transfer of preactivated OT-I CTL inhibited tumor growth in a dose-dependent manner, indicating that E.G7 was susceptible to immune effector cells. Importantly, naive OT-I T cells proliferated and elicited an anti-tumor response if they were adoptively transferred into normal or CD4-deficient mice that were then vaccinated with GM-CSF-induced bone marrow-derived OVA-pulsed APC. Collectively, these data indicate that even though naive tumor-specific T cells are present at a relatively high fraction they remain ignorant of the tumor and demonstrate that a CD8-mediated anti-tumor response can be induced by Ag-pulsed APC without CD4 T cell help.     

Thalidomide prevents bleomycin-induced pulmonary fibrosis in mice

Pulmonary fibrosis in humans can occur as a result of a large number of conditions. In idiopathic pulmonary fibrosis (IPF), pulmonary function becomes progressively compromised resulting in a high mortality rate. Currently there are no proven effective treatments for IPF. We have recently reported that IL-6 and TGF-beta(1) plays an important role in proliferation and differentiation of lung fibroblasts, and all-trans-retinoic acid (ATRA) prevented bleomycin-induced lung fibrosis through the inhibition of these cytokines. Thalidomide (Thal) has been used in the treatment of multiple myeloma through the inhibitory effect on IL-6-dependent cell growth and angiogenesis. In this study, we examined the preventive effect of Thal on bleomycin-induced pulmonary fibrosis in mice. We performed histological examinations and quantitative measurements of IL-6, TGF-beta(1), collagen type Ialpha1 (COL1A1), vascular endothelial growth factor (VEGF), angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2) in bleomycin-treated mouse lung tissues with or without the administration of Thal. Thal histologically ameliorated bleomycin-induced fibrosis in mouse lung tissues. Thal decreased the expressions of IL-6, TGF-beta(1), VEGF, Ang-1 Ang-2, and COL1A1 mRNA in mouse lung tissues. In addition, Thal inhibited angiogenesis in the lung. In vitro studies disclosed that Thal reduced 1) production of IL-6, TGF-beta(1), VEGF, Ang-1, and collagen synthesis from human lung fibroblasts, and 2) both IL-6-dependent proliferation and TGF-beta(1)-dependent transdifferentiation of the cells, which could be the mechanism underlying the preventive effect of Thal on pulmonary fibrosis. These data may provide a rationale to explore clinical use of Thal for the prevention of pulmonary fibrosis.     

CC-chemokine receptor 2 required for bleomycin-induced pulmonary fibrosis

MCP-1, which signals via the CC chemokine receptor 2 (CCR2), is induced in lung fibrosis that is accompanied by mononuclear cell recruitment and activation of lung fibroblasts. To evaluate the role of CCR2 in lung fibrosis, CCR2 knockout (ko) mice were used in a model of bleomycin-induced lung fibrosis. Wild type (wt) and ko mice were injected endotracheally with bleomycin to induce lung injury and fibrosis, and then analyzed for degree of lung fibrosis and cytokine expression. The results showed significantly reduced fibrosis in ko mice as evidenced by decreased lung type I collagen gene expression and hydroxyproline content relative to those in wt mice. Lung TNF-alpha and TGF-beta1 expression was significantly lower in ko vs. wt mice, while MCP-1 expression was unaffected. Interestingly, lung alpha-smooth muscle actin (alpha-SMA) expression, a marker for myofibroblast differentiation, was also decreased in ko mice, which was confirmed by analysis of isolated lung fibroblasts. Fibroblasts from ko mice exhibited decreased responsiveness to TGF-beta1 induced alpha-SMA expression, which was associated with reduced expression of TGF-beta receptor II (TbetaRII) and Smad3. These findings suggest that CCR2 signaling plays a key role in bleomycin-induced pulmonary fibrosis by regulating fibrogenic cytokine expression and fibroblast responsiveness to TGF-beta.     

Dual-immunohistochemistry provides little evidence for epithelial–mesenchymal transition in pulmonary fibrosis

epithelial–mesenchymal transition (EMT) has been considered to be involved in organ fibrogenesis. However, there is few direct evidence of this process in the pathophysiology of pulmonary fibrosis in vivo. Therefore, we tried to verify the involvement of this process in the development of pulmonary fibrosis. Since the co-expressions of epithelial and mesenchymal markers are thought to be a marker of EMT, we performed dual-immuunohistochemistry to assess the co-expressions of these proteins in lung tissues from bleomycin-induced pulmonary fibrosis in mice, and from patients with idiopathic pulmonary fibrosis, and nonspecific interstitial pneumonia. Double positive cells for epithelial markers including E-cadherin, T1α, or aquaporin 5, and a mesenchymal markers α-smooth muscle actin or vimentin were not found in bleomycin-induced pulmonary fibrosis in mice. Double positive cells for E-cadherin, ICAM-1, LEA, CD44v9, or SP-A and α-smooth muscle actin or vimentin were not found in lung tissues from normal lung parenchyma, idiopathic pulmonary fibrosis and nonspecific interstitial pneumonia. These results offer at least two possibilities. One is that EMT does not occur in IPF or bleomycin-induced pulmonary fibrosis in mice. Another is that EMT may occur in pulmonary fibrosis but the time during this transition in which cells express detectable levels of epithelial and mesenchymal markers is too small to be detected by double immunohistochemistry.     

Nitric oxide attenuates epithelial-mesenchymal transition in alveolar epithelial cells

Patients with interstitial lung diseases, such as idiopathic pulmonary fibrosis (IPF) and bronchopulmonary dysplasia (BPD), suffer from lung fibrosis secondary to myofibroblast-mediated excessive ECM deposition and destruction of lung architecture. Transforming growth factor (TGF)-beta1 induces epithelial-mesenchymal transition (EMT) of alveolar epithelial cells (AEC) to myofibroblasts both in vitro and in vivo. Inhaled nitric oxide (NO) attenuates ECM accumulation, enhances lung growth, and decreases alveolar myofibroblast number in experimental models. We therefore hypothesized that NO attenuates TGF-beta1-induced EMT in cultured AEC. Studies of the capacity for endogenous NO production in AEC revealed that endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) are expressed and active in AEC. Total NOS activity was 1.3 pmol x mg protein(-1) x min(-1) with 67% derived from eNOS. TGF-beta1 (50 pM) suppressed eNOS expression by more than 60% and activity by 83% but did not affect iNOS expression or activity. Inhibition of endogenous NOS with l-NAME led to spontaneous EMT, manifested by increased alpha-smooth muscle actin (alpha-SMA) expression and a fibroblast-like morphology. Provision of exogenous NO to TGF-beta1-treated AEC decreased stress fiber-associated alpha-SMA expression and decreased collagen I expression by 80%. NO-treated AEC also retained an epithelial morphology and expressed increased lamellar protein, E-cadherin, and pro-surfactant protein B compared with those treated with TGF-beta alone. These findings indicate that NO serves a critical role in preserving an epithelial phenotype and in attenuating EMT in AEC. NO-mediated regulation of AEC fate may have important implications in the pathophysiology and treatment of diseases such as IPF and BPD.     

Transforming growth factor (TGF)-beta1 stimulates pulmonary fibrosis and inflammation via a Bax-dependent, bid-activated pathway that involves matrix metalloproteinase-12

Fibrosis, apoptosis, and the exaggerated production of transforming growth factor (TGF)-beta(1) are juxtaposed in a variety of pulmonary diseases including the interstitial lung diseases and asthma. In these disorders, the relationships between these responses are not well defined. In addition, the apoptosis pathways that contribute to these responses and the mechanism(s) of their contribution have not been described. We hypothesized that BH3 domain-only protein-induced apoptosis plays an important role in the pathogenesis of TGF-beta(1)-induced pulmonary responses. To test this hypothesis, we characterized the effects of transgenic TGF-beta(1) in mice with wild type (WT) and null Bax loci. To investigate the mechanisms of Bax activation and its effector functions, we also compared the effects of TGF-beta(1) in mice with WT and null Bid and matrix metalloproteinase (MMP)-12 loci, respectively. These studies demonstrate that TGF-beta(1) is a potent stimulator of Bax, Bid, and MMP-12. The studies also demonstrate that Bax and Bid play key roles in the pathogenesis of TGF-beta(1)-induced inflammation, fibrosis, and apoptosis; that TGF-beta(1) stimulates MMP-12, TIMP-1, and cathepsins and inhibits MMP-9 and p21 via Bax- and Bid-dependent mechanisms; and that TGF-beta(1)-stimulated pulmonary fibrosis is ameliorated in MMP-12-deficient animals. Finally, they demonstrate that Bax, Bid, and MMP-12 play similar roles in bleomycin-induced fibrosis, thereby highlighting the importance of this Bid-activated, Bax-mediated pathway and downstream MMP-12 in a variety of fibrogenic settings.     

TGF-beta1 在不同类型特发性间质性肺炎患者肺组织中的表达及其意义

目的:探讨TGF-β1在不同类型特发性间质性肺炎患者肺组织中的表达及其意义。方法:选取2010年2月至2013年12月在我院就诊的72例经支气管镜肺活检的不同类型的特发性间质性肺炎患者的组织标本,对其转化生长因子-β1表达程度进行评定。结果:寻常型(普通型)、非特异性、脱屑性、急性等ⅡP、呼吸性细支气管炎并间质性肺疾病以及隐原性机化性肺炎患者肺组织中TGF-β1表达强度评分均显著高于对照组;脱屑性ⅡP和呼吸性细支气管炎并间质性肺疾病患者肺组织中TGF-β1表达强度评分均显著高于其他类型ⅡP患者;非特异性ⅡP、急性ⅡP、淋巴细胞性ⅡP以及隐原性机化性肺炎组患者肺组织中TGF-β1表达强度评分均显著低于寻常型(普通型)ⅡP组;隐原性机化性肺炎、淋巴细胞性ⅡP组患者肺组织中TGF-β1表达强度评分分别为(0.93±0.34)分、(0.82±0.27)分,显著低于急性ⅡP组患者的(1.64±0.05)分。差异均有统计学意义(P<0.05)。结论:TGF-β1表达过度可能是ⅡP患者的重要特征,在肺纤维化的过程中发挥着重要作用,但在不同类型ⅡP中的作用机制并不相同。     

Mechanisms of oncostatin M-induced pulmonary inflammation and fibrosis

Oncostatin M (OSM), an IL-6 family cytokine, has been implicated in a number of biological processes including the induction of inflammation and the modulation of extracellular matrix. In this study, we demonstrate that OSM is up-regulated in the bronchoalveolar lavage fluid of patients with idiopathic pulmonary fibrosis and scleroderma, and investigate the pathological consequences of excess OSM in the lungs. Delivery of OSM to the lungs of mice results in a significant recruitment of inflammatory cells, as well as a dose-dependent increase in collagen deposition in the lungs, with pathological correlates to characteristic human interstitial lung disease. To better understand the relationship between OSM-induced inflammation and OSM-induced fibrosis, we used genetically modified mice and show that the fibrotic response is largely independent of B and T lymphocytes, eosinophils, and mast cells. We further explored the mechanisms of OSM-induced inflammation and fibrosis using both protein and genomic array approaches, generating a "fibrotic footprint" for OSM that shows modulation of various matrix metalloproteinases, extracellular matrix components, and cytokines previously implicated in fibrosis. In particular, although the IL-4/IL-13 and TGF-beta pathways have been shown to be important and intertwined of fibrosis, we show that OSM is capable of inducing lung fibrosis independently of these pathways. The demonstration that OSM is a potent mediator of lung inflammation and extracellular matrix accumulation, combined with the up-regulation observed in patients with pulmonary fibrosis, may provide a rationale for therapeutically targeting OSM in human disease.     

Gene transfer of hepatocyte growth factor by electroporation reduces bleomycin-induced lung fibrosis

Abnormal alveolar wound repair contributes to the development of pulmonary fibrosis after lung injury. Hepatocyte growth factor (HGF) is a potent mitogenic factor for alveolar epithelial cells and may therefore improve alveolar epithelial repair in vitro and in vivo. We hypothesized that HGF could increase alveolar epithelial repair in vitro and improve pulmonary fibrosis in vivo. Alveolar wound repair in vitro was determined using an epithelial wound repair model with HGF-transfected A549 alveolar epithelial cells. Electroporation-mediated, nonviral gene transfer of HGF in vivo was performed 7 days after bleomycin-induced lung injury in the rat. Alveolar epithelial repair in vitro was increased after transfection of wounded epithelial monolayers with a plasmid encoding human HGF, pCikhHGF [human HGF (hHGF) gene expressed from the cytomegalovirus (CMV) immediate-early promoter and enhancer] compared with medium control. Electroporation-mediated in vivo HGF gene transfer using pCikhHGF 7 days after intratracheal bleomycin reduced pulmonary fibrosis as assessed by histology and hydroxyproline determination 14 days after bleomycin compared with controls treated with the same vector not containing the HGF sequence (pCik). Lung epithelial cell proliferation was increased and apoptosis reduced in hHGF-treated lungs compared with controls, suggesting increased alveolar epithelial repair in vivo. In addition, profibrotic transforming growth factor-beta1 (TGF-beta1) was decreased in hHGF-treated lungs, indicating an involvement of TGF-beta1 in hHGF-induced reduction of lung fibrosis. In conclusion, electroporation-mediated gene transfer of hHGF decreases bleomycin-induced pulmonary fibrosis, possibly by increasing alveolar epithelial cell proliferation and reducing apoptosis, resulting in improved alveolar wound repair.