Elevated Tenascin-C Levels in Bronchoalveolar Lavage Fluid of Patients with Sarcoidosis

Background

Sarcoidosis is a disease characterized by granulomatous lesions involving multiple organ systems. The etiology of sarcoidosis remains unknown, and reliable biomarkers have not been identified. Tenascin-C is an extracellular matrix molecule expressed during wound healing in various tissues. The present study aimed to investigate the role of tenascin-C in sarcoidosis.

Methods

Enzyme-linked immunosorbent assays were used to measure tenascin-C levels in serum and bronchoalveolar lavage fluid (BALF) from 31 patients with sarcoidosis and 15 healthy individuals. Relationships between tenascin-C concentrations in BALF and serum samples and clinical parameters in patients with sarcoidosis were evaluated.

Results

BALF tenascin-C levels were significantly higher in patients with sarcoidosis than in healthy individuals, but serum levels were no different. BALF tenascin-C levels showed positive correlations with serum lactic dehydrogenase levels and the ratio of lymphocytes in BALF. BALF tenascin-C levels were also higher in patients with parenchymal infiltration on chest radiographs than in those without.

Conclusions

The present results demonstrated that the BALF tenascin-C level was correlated with pulmonary infiltrates on chest radiographs in patients with sarcoidosis. Although measurement of serum tenascin-C levels has a limited role and measurement of BALF tenascin-C levels might be impractical, tenascin-C in the lung might play a role in the pathogenesis of sarcoidosis. Further studies are necessary to determine the role of BALF tenascin-C in sarcoidosis.     

Isoniazid Levels in the Bronchoalveolar Lavage Fluid of Patients with Pulmonary Tuberculosis

Isoniazid (INH) is one of the most important first line drugs in the treatment of tuberculosis. We utilized high performance liquid chromatography with a hydrazone extraction technique to measure INH in bronchoalveolar lavage (BAL) fluid specimens from six patients with active pulmonary tuberculosis. We found BAL fluid INH levels to be similar to 2-h peak serum levels. The concentration of INH in BAL fluid from lobes with infiltrate was similar to the concentration of INH in BAL fluid from lobes without infiltrate (0.062 μg/ml and 0.073 μg/ml, respectively). After adjusting for protein concentration in the BAL fluid, INH levels in lobes with infiltrate were threefold lower than in lobes without infiltrate. The correlation between the concentration of INH in serum and BAL fluid approached significance after correcting for protein (lobes with infiltrate, r ²= 0.60 (p= 0.07); lobes without infiltrate, r ²= 0.50 (p= 0.12)). INH penetrates into bronchoalveolar fluid, and concentrations of INH in the BAL fluid suggest that assessment of the INH serum concentration is adequate to evaluate bioavailability of the drug in patients with pulmonary tuberculosis. Accepted for publication: 26 June 1997     

Elevated Levels of Type II Soluble Tumor Necrosis Factor Receptors in the Bronchoalveolar Lavage Fluids of Patients with Sarcoidosis

Since tumor necrosis factor (TNF) is known to be involved in granuloma formation in sarcoidosis, and soluble TNF receptors (sTNF-Rs) inhibit TNF action in vivo, we evaluated the levels of sTNF-Rs in the bronchoalveolar lavage fluids (BALF) of 31 subjects using an enzyme-linked immunosorbent assay. Our group consisted of 13 patients with sarcoidosis (7 sarcoidosis patients who received no treatment and 6 who received corticosteroid therapy) and 18 control subjects (11 healthy nonsmokers and 7 asymptomatic smokers). Type II (75-kDa), but not type I (55 kDa) sTNF-R in BALF was elevated significantly in patients with sarcoidosis compared with the healthy nonsmokers (type I: 126.7 ± 17.6 pg/ml BALF vs 79.4 ± 16.5 pg/ml BALF, p > 0.05; type II: 98.3 ± 27.8 pg/ml BALF vs 26.7 ± 4.9 pg/ml BALF, p < 0.05). Although levels of type I sTNF-R in BALF from sarcoidosis patients were not correlated with any cellular profiles of BALF, concentrations of type II correlated significantly with the numbers of lymphocytes in BALF. We concluded that sTNF-R is a normal constituent of the epithelial lining fluids and that levels of type II sTNF-R are elevated significantly in the BALF from individuals with sarcoidosis. This suggests that sTNF-Rs may influence the local bioactivity of TNF and may also contribute to the pathogenesis of sarcoidosis. Accepted for publication: 7 November 1996     

Mast Cells and Lysozyme Positive Macrophages in Bronchoalveolar Lavage from Patients with Sarcoidosis

ABSTRACT In the deteriorating group of sarcoidosis patients, progress towards pulmonary fibrosis is a major problem. In order to benefit from corticosteroids, it is important for the treatment to start early. We studied a group of 45 patients with sarcoidosis. Most of them were newly detected patients and none were under or had currently received corticosteroid therapy. The patients were followed for at least six months. We found that increased amounts of polymorphonuclear neutrophils (PMN) or lysozyme-positive macrophages (Lys⁺MF) and mast cells (MC) in bronchoalveolar lavage (BAL) could implicate a bad prognosis.     

乌司他丁对体外循环患者肺灌洗液中炎性因子和肺功能的影响

目的:探讨丝氨酸蛋白酶抑制剂乌司他丁对体外循环下心脏直视手术患者肺灌洗液中炎性因子和肺功能的影响。方法:将30例拟行瓣膜置换术患者随机分为乌司他丁组和对照组,各15例。左心室射血分数<0．45、左心室舒张末直径>70 mm、心胸比率>0．7、血流动力学不稳定以及肝、肾功能障碍和急症手术的患者不选。乌司他丁组:在劈胸骨和升主动脉开放前分别给予乌司他丁0．6 KIU／kg;对照组:与乌司他丁组同时点只给予等量生理盐水。在术前和升主动脉开放后30分钟进行右肺中叶灌洗,采集肺灌洗液,用酶联免疫法测定中性粒细胞弹性蛋白酶和白细胞介素-8浓度,并在术前、升主动脉开放后30分钟、2小时和6小时记录呼气末二氧化碳(PETCO2)、采集桡动脉血行血气分析,计算肺功能指标:肺泡-动脉氧分压差[P(A-a)DO2]、氧合指数(PaO2／FiO2)和死腔率(VD／VT)。结果:①对照组肺灌洗液中白细胞介素-8浓度在升主动脉开放后30分钟升高,与术前比较差异有统计学意义(P< 0．05);乌司他丁组肺灌洗液中白细胞介素-8浓度在升主动脉开放后30分钟也较术前升高,但无显著差异(P>0．05),但乌司他丁组低于对照组同时点水平,差异有统计学意义(P<0．05);②对照组体外循环后各时点P(A-a)、DO2、VD／VT均较术前升高。P(A-a)DO2在升主动脉开放后30分钟、升主动脉开放后2小时升高与术前相比有统计学意义(P<0．05), VD／VT在升主动脉开放后30分钟升高达峰值(P<0．05);乌司他丁组在体外循环后各时点P(A-a)DO2、VD／VT也较术前升高,但无显著差异(P>0．05),在升主动脉开放后30分钟均低于对照组,差异有统计学意义(P<0．05);③对照组PaO2／FiO2体外循环后各时点与术前相比均降低,在升主动脉开放后30分钟降到最低值(P<0．05);乌司他丁组PaO2／FiO2在体外循环后各时点也降低,但无显著差异(P>0．05),在升主动脉开放后30分钟显著高于对照组(P<0．05)。结论:体外循环下心脏直视手术可致肺功能损伤,乌司他丁可抑制肺内炎性因子白细胞介素-8和弹性蛋白酶释放而具有肺保护作用。     

Level of Nitric Oxide in Bronchoalveolar Lavage Fluid of Asthmatic Mice Model

Background: Asthma is a chronic inflammatory disease with multifactorial and complicated mechanisms. Elevated level of exhaled Nitric Oxide (NO) in asthma and other inflammatory lung diseases has led to many studies examining NO as a potential marker of airway inflammation. Objective: This study was designed to determine the level of NO in Bronchoalveolar Lavage (BAL) fluid during early and late stages of asthmatic attack in mouse model.   Methods: In this study male BALB/c mice were used. The level of NO was determined in BAL fluid of asthmatic mice five minutes, six and sixteen hours after challenge with methacholine, as irritant and smoke and 5% ovalbumin as allergens, using colorimetric assay.   Results: The level of NO increased upon exposure to all three irritants used in this study (52.3   μM for smoke and 49.5 μ   Mfor methacholine) as compared to 22.8 μM for the baseline. Our results showed that NO levels were increased during early phase of asthmatic condition and reached to its maximum level after six hours and decreased at the late stage of asthma (16hrs) possibly by activating a feedback regulatory loop. In addition, high level of NO led to the hypertrophy of smooth muscle that can account for the pathological changes associated with asthma.   Conclusion: Thus, NO is an inflammatory marker in asthma and its measurement, as a non-invasive method during asthmatic attack is suggested. A careful development of specific inhibitors for iNOS enzyme during asthmatic attack is also necessary.     

Changes of Soluble ICAM-1 Levels in Serum and Bronchoalveolar Lavage Fluid from Patients with Atopic Bronchial Asthma After Allergen Challenge

Leukocyte-endothelial cell interaction is essential for leukocyte infiltration into inflammatory sites. Initiation of adhesion is through the up-regulated expression of adhesion molecules in the endothelium or epithelium and the activation of adhesion molecules on leukocytes. To our knowledge, there have been few reports concerning soluble intercellular adhesion molecule-1 (s1CAM-1) in patients with atopic bronchial asthma after allergen challenge. If the levels of s1CAM-1 vary between bronchial asthma patients and normal controls, this variance would be useful to assess the state of this disease. Therefore, we measured the levels of s1CAM-1 in sera from 17 patients with atopic bronchial asthma and normal control subjects. Levels of s1CAM-1 in sera from bronchial asthma patients in prechallenge conditions were higher than in normal control subjects. Levels of s1CAM-1 in sera from bronchial asthma patients 8 hr after challenge were higher than those in sera obtained during prechallenge periods. s1CAM-1 levels in bronchoalveolar lavage (BAL) fluids from bronchial asthma patients 8 hr after challenge were higher than at 30 min after challenge. These results suggest that higher levels of s1CAM-1 in sera and BAL fluids reflect the up-regulation of ICAM-1 expression in allergic bronchial asthma and these high levels may contribute to the pathogenesis of atopic bronchial asthma.     

肺孢子虫肺炎大鼠支气管肺泡灌洗液酶含量变化及大蒜素治疗对其影响

目的 研究卡氏肺孢子虫肺炎(PCP)大鼠支气管肺泡灌洗液(BALF)酶含量变化以及大蒜素治疗对其影响。 方法 用地塞米松连续肌肉注射Wistar大鼠， 诱导建立PCP大鼠模型。诱导第3、6、9周分别于后腿肌肉深部注射大蒜素治疗（10 mg/kg， 1次/d， 连续5 d）。同时设甲氧苄氨嘧啶(TMP)?鄄磺胺甲基异噁唑(SMZ)治疗对照组(SMZ/TMP组)、 PCP模型对照组和空白对照组。 于最后1次治疗3 d后处死大鼠， 无菌收集BALF， 测定谷草转氨酶(AST)、 谷丙转氨酶(ALT)、 胆碱脂酶(CHE)、 碱性磷酸酶(ALP)、 乳酸脱氢酶(LDH)、 肌酸激酶(CK)及其同工酶(CKMB)、 a?鄄羟丁酸脱氢酶(HBDH)、 a?鄄L?鄄岩藻糖甘酶(AFU)、 5′-核苷酸酶(5′NT)及腺苷脱氨酶(ADA)含量。 结果 PCP模型组ALP含量［（573.41 ± 350.63） U/L］显著高于空白对照组［（210.56 ± 114.41） U/L］（q=4.682， P<0.01）、 大蒜素治疗组［（392.07 ± 217.57） U/L］（q=3.851， P<0.05）以及SMZ/TMP组［（325.21 ± 180.65） U/L］（q=4.380， P<0.01）。 CK、 CKMB及5′NT含量， PCP模型对照组［依次为948.94 ± 403.43、 489.47 ± 254.46及（6.76 ± 3.11） U/L］显著高于空白对照组［426.22 ± 319.00、 213.33 ± 144.54及（3.22 ± 1.20） U/L］（q=4.696， 3.784， 3.812， P<0.05）。AST、 ALT、 CHE、 LDH、 HBDH、 AFU及ADA含量， 4组之间差异均无统计学意义(F=1.852， 0.958， 2.470， 1.423， 1.178， 1.342， 0.611， P>0.05)。 结论 PCP大鼠BALF中ALP、 CK、 CKMB及5′NT含量显著升高, 大蒜素治疗可使ALP含量显著降低。     

从大鼠肺灌洗液中分离纯化培养卡氏肺孢子虫

目的 建立卡氏肺孢子虫（Pneumocystis carinii, P.c）纯培养株。 方法　 从肺孢子虫肺炎（PCP）大鼠模型离体肺脏的灌洗液中分离P.c，用改良IMDM培养基进行体外培养；以四胺银染色计数法观察虫体增殖情况；用PCR扩增培养物中P.c线粒体大亚基rRNA基因，进行基因鉴定；用透射电镜观察培养虫体的超微结构。 结果 用添加ｓ-腺苷甲硫氨酸（SAM）等辅助剂的IMDM培养基从8只PCP大鼠的肺灌洗液中分离出5个P.c纯培养株（P.c Rat1~5）。分离培养的P.c可进行冷冻保存和复苏培养。培养72 h的P.c包囊可增殖19~22倍。形态、超微结构及基因序列分析证实从大鼠肺灌洗液中分离出的培养物是大鼠源性肺孢子虫。 结论 从大鼠肺灌洗液中分离培养出5株大鼠源卡氏肺孢子虫。     

Th17细胞及其细胞因子白细胞介素-17在粉尘螨致敏哮喘小鼠中的变化

目的探讨Th17细胞及其细胞因子白细胞介素-17(IL-17)在粉尘螨致敏哮喘小鼠中的变化及其意义。方法 20只BALB/c小鼠随机均分为哮喘组和健康对照组。哮喘组小鼠于第0、7和14天每鼠腹腔注射200μl致敏液[含粉尘螨粗浸液提取物50μg,Al(OH)32 mg]致敏。末次免疫后1周采用粉尘螨粗浸液提取物连续进行滴鼻激发,每天1次,每次50μg,激发7次,健康对照组给予等体积的PBS[含Al(OH)32 mg]处理。末次激发后24 h内处死小鼠,取血清和肺泡灌洗液,无菌取脾脏。ELISA检测血清中Ig G1、Ig E和肺泡灌洗液中IL-17的含量;流式细胞仪检测脾脏中Th17细胞百分率。结果哮喘组小鼠血清中Ig G1和Ig E水平分别为(0.10±0.01)pg/ml和(1.15±0.10)pg/ml,高于健康对照组的(0.06±0.01)pg/ml和(0.04±0.01)pg/ml(P0.05);哮喘组小鼠肺泡灌洗液IL-17水平(85.13±2.36)pg/ml高于健康对照组(48.27±4.14)pg/ml(P0.01);哮喘组小鼠脾脏Th17细胞百分率(5.19±0.68)%高于健康对照组(0.95±0.19)%(P0.01),且脾脏Th17细胞百分率与肺泡灌洗液中IL-17水平呈正相关(r=0.851,P0.01)。结论粉尘螨致敏哮喘小鼠较健康小鼠肺泡灌洗液IL-17水平和脾脏Th17细胞数量均升高。     

The Validity of Induced Sputum and Bronchoalveolar Lavage in Childhood Asthma

There are a number of useful direct airway sampling procedures to help diagnose and monitor asthma in patients. However, non-invasive techniques are the ideal, especially in children, given the necessity of safe and repeatable measurements to monitor treatment efficacy and disease progression. Bronchoalveolar lavage (BAL) may be too invasive for clinical use in children, while questions still surround the utility of induced sputum (IS). More novel techniques, such as fractional exhaled nitric oxide (FE_NO) and exhaled breath condensate (EBC), are still unproven. Eosinophilic airway inflammation is a major feature of childhood asthma, and it has been revealed as a major treatment target with inhaled corticosteroids. Moreover, treatment protocols governed by sputum eosinophil counts may be more efficacious – by reducing the frequency and severity of exacerbations – than treatment based on clinical symptoms and other traditional objective measures of lung function. The selection of an appropriate airway inflammation monitoring technique must take everything into consideration, including safety, reproducibility, repeatability, sensitivity to treatment, and the overall clinical/research goals.     

Utility of Bronchoalveolar Lavage and Transbronchial Biopsy in Patients with Hypersensitivity Pneumonitis

Introduction

Making the diagnosis of HP is challenging due to a lack of consensus criteria and variability of both pathologic and radiographic findings. The purpose of this retrospective study was to determine the diagnostic utility of the combination of BAL lymphocyte count and TBBX in patients with HP.

Methods

We conducted a retrospective cohort study of all patients with a MDD diagnosis of HP at a single center.

Results

155 patients were included in the study. 49% of patients who underwent BAL had a lymphocyte count?>?20, 42% had a lymphocyte count?>?30, and 34% had lymphocyte count?>?40%. The median BAL lymphocyte count was higher in inflammatory HP compared to fibrotic HP. The addition of TBBX to BAL significantly increased the diagnostic yield regardless of the BAL lymphocyte cutoff used. The yield of bronchoscopy with TBBX and BAL when a lymphocyte count?>?40% was used as a cutoff was 52%.

Conclusions

Our study suggests that the combination of TBBX with BAL significantly increases the likelihood that the procedure will provide adequate additional information to allow a confident MDD diagnosis of HP and may reduce the need for SLB in the diagnostic workup of HP.

     

Different Expression of Endothelin in the Bronchoalveolar Lavage in Patients with Pulmonary Diseases

Endothelin (ET) is a broncho- and vasoconstrictive cytokine, but it also possesses proinflammatory and mitogenic activity. It is suggested to be involved in the pathogenesis of fibrotic lung diseases. We analyzed the concentration of ET 1 in the bronchoalveolar lavage (BAL) fluid in 95 patients with different lung diseases, among them 41 patients with interstitial lung diseases (13 fibrosing alveolitis in systemic sclerosis (FASS), 9 idiopathic pulmonary fibrosis (IFP), 8 sarcoidosis (S), 6 occupational lung disease (OLD), 5 other alveolitidies A), 27 patients with pneumonia, and 8 patients with chronic obstructive pulmonary disease (COPD). A heterogeneous group of 19 patients served as controls. The median ET concentration was 3.3 pg/ml. Significantly higher concentration was found in patients with FASS (5.8 pg/ml), IPF (5.0 pg/ml), and S (5.1 pg/ml) compared with OLD (2.8 pg/ml), A (1.9 pg/ml), COPD (1.5 pg/ml), and the control group (2.5 pg/ml). In pneumonia, the elevated ET concentration (4.1 pg/ml) was accompanied by a high alveolocapillary leakage. When normalized to BAL albumin concentration, only FASS presented with significantly elevated ET/albumin in the BAL compared with the control group (134.5 vs. 56.l pg/mg, p < 0.05). There were no correlations between ET and BAL differential cell count or pulmonary function tests. In current smokers, ET in BALF was significantly higher compared with non- or ex-smokers (3.9 vs. 2.0 pg/ml, p < 0.01), but not so the ET/albumin ratio (65.0 vs. 62.5 pg/mg). In summary, ET in the BAL is differentially expressed in distinct inflammatory and interstitial lung disease. Consistently high concentrations are found in FASS and elevated ET concentration could be discussed in IPF, sarcoidosis, and pneumonia. ET concentration in BAL is influenced by current smoking habits.     

Tissue and Bronchoalveolar Lavage Biomarkers in Idiopathic Pulmonary Fibrosis Patients on Pirfenidone

Background

Pirfenidone is a novel anti-fibrotic agent in idiopathic pulmonary fibrosis with proven clinical benefit. Better human tissue models to demonstrate the immunomodulatory and anti-fibrotic effect of pirfenidone are required.

Objectives

The purpose of the study was to use transbronchial lung cryobiopsy (TBLC), a novel technique which provides substantial tissue samples, and a large panel of biomarkers to temporally assess disease activity and response to pirfenidone therapy.

Methods

Thirteen patients with confirmed idiopathic pulmonary fibrosis (IPF) underwent full physiological and radiological assessment at diagnosis and after 6-month pirfenidone therapy. They underwent assessment for a wide range of potential serum and bronchoalveolar lavage biomarkers of disease activity. Finally, they underwent TBLC before and after treatment. Tissue samples were assessed for numbers of fibroblast foci, for Ki-67, a marker of tissue proliferation and caspase-3, a marker of tissue apoptosis.

Results

All patients completed treatment and investigations without significant incident. There was no significant fall in number of fibroblast foci per unit tissue volume after treatment (pre-treatment: 0.14/mm² vs. post-treatment 0.08/mm², p?=?0.1). Likewise, there was no significant change in other markers of tissue proliferation, Ki-67 or Caspase-3 with pirfenidone treatment. We found an increase in three bronchoalveolar lavage angiogenesis cytokines, Placental Growth Factor, Vascular Endothelial Growth Factor-A, and basic Fibroblast Growth Factor, two anti-inflammatory cytokines Interleukin-10 and Interleukin-4 and Surfactant Protein-D.

Conclusions

TBLC offers a unique opportunity to potentially assess the course of disease activity and response to novel anti-fibrotic activity in IPF.