INTRODUCTION

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION REFERENCES

Lung transplantation has become an accepted therapy for patients with end-stage lung diseases. However, certain long-term problems exist, with a 5-yr survival rate of only 40 to 45% (1). Long-term survival has been compromised mainly by obliterative bronchiolitis (OB). This complication denotes a histopathological process in which the terminal bronchioles show evidence of scarring progressing to eventual total occlusion of the bronchiolar lumen (2). In addition, a clinical entity of functional deterioration in graft performance known as obliterative bronchiolitis syndrome (OBS) has been defined. The diagnosis of OBS is based upon a 20% or greater decline from the average of the two highest postoperative consecutive FEV₁ measurements (3). OBS occurs in 20 to 40% of lung recipients and is characterized by dyspnea and progressive obstructive lung disease (4).

The etiology of OB remains to be identified. Among several possibilities of mechanisms hypothesized to play a role include infection (5), rejection (6), surgical interruption of bronchial circulation and lymphatic drainage, and denervation (7). Recurrent episodes of acute rejection have been identified as a risk factor for the development of OB (4), although chronic allograft rejection has been inferred as the most likely cause of OB (8).

Obliterative bronchiolitis occurs unpredictably, is poorly detected in the preclinical stage, and usually cannot be treated successfully once it is clinically apparent. For this reason, OB constitutes the most important threat to the long-term survival of lung transplant recipients. To date, the gold standard method to follow graft function is based upon histologic grading (2, 9), an invasive technique requiring transbronchial biopsy. Moreover, diagnostic certainty may be compromised by tissue sampling error related to histopathological nonuniformity of the process (9). Therefore, an important need exists for more effective and preferably less invasive approaches to monitor graft function. In view of the latter, and given the paucity of data in the literature related to lung mechanics after double-lung (DL) transplantation in humans, this study examined lung mechanics in a population of patients after DL transplantation, with emphasis on detecting the perturbations in small airways function that parallel the structural abnormalities found in OB.

	METHODS

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION REFERENCES

Lung Transplant Subjects

Subjects were recruited from an adult patient population who underwent DL transplantation at the Hospital of the University of Pennsylvania (HUP), from March 1993 to April 1997. During this period 75 such procedures were performed; 51 survived at the time of the study. Thirty-four subjects who fulfilled the inclusion criteria were approached with the goals of the study and 15 agreed to participate. The 15 subjects in the participating group did not differ significantly from the 19 subjects in the nonparticipating group in age, height, and gender. The study was approved by the institutional review board of Children's Hospital of Philadelphia (CHOP). Informed consent was obtained from each subject.

Inclusion criteria required that patients were in stable clinical condition at the time of testing with no hospital admissions during the 12 mo prior to study. Accordingly, the patients had no evidence of dyspnea, hypoxemia, hypercarbia, acute upper or lower respiratory tract infection, a positive chest X-ray, or any evidence of bronchial stenosis in the allograft. All patients continued with their usual medications on the day of study.

The investigators performing the tests were blinded to all patients' previous medical history except for age, and dates of transplant and last admission. Other clinical information became available only after completion of the study. Clinical information was provided by their primary physician, and included medical records, chest X-rays, bronchoscopy results, and transbronchial biopsy reports. According to this information, patients were classified into the following two groups:

Double lung recipient in stable condition (DL-S): This group of 11 patients had no evidence of significant infection or OBS at time of testing. Nine members had transbronchial biopsies in the year prior to the study. None of these biopsies showed significant rejection, defined as histologic acute rejection of greater than minimal grade ( 2), or as active OB as stated by the Lung Rejection Study Group (2). Significant infection was defined as histologic changes consistent with viral, fungal, or bacterial infection, or by the presence of such organisms in the sputum or bronchoalveolar lavage (BAL) that required treatment. Asymptomatic cytomegalovirus (CMV) was defined as the presence of CMV by early antigen or positive culture from any body site in the absence of symptoms.

Double lung recipient with obliterative bronchiolitis syndrome (DL-OBS): OBS was defined by a drop in 20% from the average of two best consecutive post-transplant values of FEV₁ (3). This group consisted of four subjects. Two had transbronchial biopsies; one showed evidence of OB and the second subject did not show significant rejection. None had evidence of significant infection at the time of the study.

Control Subjects

This group comprised 15 volunteers from hospital personnel. Control subjects were matched as closely as possible for age, gender, and height relative to the transplant subjects. All were nonsmokers and free of any acute or chronic lung disease.

Pulmonary Function Tests

All pulmonary function tests (PFT) and lung mechanics measurements were performed at CHOP. These included lung volume subdivisions, forced flow parameters, and specific airway conductance (sGAW). These were measured by body plethysmograph (BP). We also measured functional residual capacity by helium dilution (FRC_He), and expressed the ratio FRC_He/FRC_BP to quantify air trapping. Arterial oxygen saturation (Sa_O2) and expired end-tidal CO₂ tension (PET_CO2) were measured with a pulse oximeter and capnometer while patients were at rest. An individual's PFT results were considered normal if they were within ± 2 SD from reference values (10- 12).

We performed the single-breath nitrogen test according to a standard method (13), to assess for the presence of small airways dysfunction manifested by inhomogeneity of N₂ washout from the lungs and reflected by an elevated slope of phase 3 (N₂SP3) (14). Normal values of the N₂SP3 were derived from the control group and available data (15, 16).

Pulmonary Mechanics

Static lung compliance curve: The method for esophageal manometry is well established in the literature (17). Briefly, a thin-walled, 10-cm-long latex balloon was attached over the perforated end of a 1.9-mm polyethylene catheter. The balloon catheter was passed transnasally after topical anesthesia and was advanced 40 to 45 cm (18). The proximal end of the catheter was attached to a glass syringe and to a pressure transducer and 0.5 ml of air was introduced into the evacuated balloon. Recording fidelity was established by adjusting the catheter placement until cardiac oscillations were minimal and pressure swings with inspiration were most negative.

To obtain the static lung pressure-volume curve we used methods previously described (19). Briefly, after placing the balloon, subjects performed three full inhalations and exhalations. These were followed by a maximal inspiration to TLC which was transiently held by a manually controlled shutter. A passive exhalation from TLC to FRC was performed by periodically opening the shutter to allow airflow. Five to seven maneuvers were performed by each subject. At each plateau volume, the difference of mouth pressure (Pm) and esophageal pressure (Pes) was the transpulmonary pressure (Pst,L). Signals were collected and analyzed by a computerized data acquisition system. The static lung pressure-volume curve was derived after plotting lung volume (as percentage of predicted TLC) versus Pst,L and fitting an exponential curve to the data points (20). The following data were obtained:

(a) Lung chord compliance (Cst,L); calculated as the volume change over Pst (l) change between FRC and FRC + 0.5 liter.

(b) Specific chord compliance (SCst,L); chord compliance normalized for the subject's FRC (Cst,L/FRC).

(c) Chord compliance normalized for the subject's TLC (Cst,L/TLC).

(d) Lung elastic recoil pressure at 90% TLC (Pst,L 90% TLC)

(e) The exponential constant (K) reflecting the shape of the pressure-volume curve.

Dynamic lung compliance (Cdyn,L): Subjects breathed at tidal volume through a mouthpiece at increasing frequencies to 1.5 Hz. Division of inspiratory volume by Pst (l) change between successive zero-flow points for each breath yielded the Cdyn,L for any given frequency. Dynamic compliance at tidal breathing (Cdyn,L), at 0.75 Hz, and the ratio of Cdyn,L/Cst,L at tidal breathing were assessed (21, 22).

Upstream resistance (R_US): This test is considered a sensitive technique to detect patency of the small airways (23). After the removal of the esophageal balloon, subjects performed 3 to 5 maximal expiratory flow maneuvers from TLC. Expiratory flows were plotted on a percentage of predicted TLC axis along with the static lung pressure-volume curve obtained previously. Below 60% vital capacity, at any given volume, lung recoil pressure divided by the corresponding flow yielded the upstream resistance of the small airways (Figure 1).

View larger version (18K): [in this window] [in a new window]   Figure 1.   A representative graph of a DL-S subject (dotted line) and a matched control subject (continuous line) describing the RUS measurement. RUS was calculated by obtaining multiple pairs of isovolume lung recoil pressures and airflow measurements from the effort independent flow region (< 60% VC). These points were plotted on Flow/Pst,L axes. The slope of linear regression was calculated as RUS.

Statistical Analysis

Pulmonary function test data were expressed as percentage of predicted of normal using previously established normative data (10). Means and SD were calculated for demographic data, PFT, and lung mechanics variables for each group studied. An unpaired t test was used to compare each variable between DL-S and control subjects, and between DL-OBS and control subjects. Pearson correlations were calculated between N₂SP3 and various PFT and R_US measurements, and between FEV₁ percentage of predicted and compliance measurements. Statistical significance was declared at a p value < 0.05.

	RESULTS

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION REFERENCES

A total of 15 patients who underwent DL transplantation participated in the study. The indications for DL transplantation were: chronic obstructive pulmonary disease (COPD, n = 7), pulmonary hypertension (n = 3), cystic fibrosis (n = 2), -1 antitrypsin deficiency (n = 2), and OB (n = 1).

Of the 15 transplant patients, 11 were classified as the DL-S group. Six of those were asymptomatic for CMV. Four were classified as the DL-OBS group, and of them three were asymptomatic for CMV. The mean time from transplantation to study was 31.4 ± 11 mo in the DL-S group, and 36.2 ± 6.8 mo in the DL-OBS (p = NS).

Demographic data on the 11 DL-S, 4 DL-OBS patients, and 15 control subjects are presented in Table 1. The mean age of the DL-S group was 47.2 ± 12.0 yr, significantly higher than control subjects 32.9 ± 5.6 yr (p < 0.001), and their donors 27.9 ± 10.8 yr (p < 0.0001). The mean DL-S height was 172 ± 5 cm and was similar to controls' and donors' height (174 ± 10 cm and 174 ± 5 cm, respectively).

The mean DL-OBS age was 37.0 ± 14.0 yr, similar to control subjects' 32.9 ± 5.6 yr, and to their donors' 23.7 ± 8.6 yr (p = NS). The mean DL-OBS height was 171 ± 4 cm and was similar to controls' and donors' height (174 ± 10 cm and 172 ± 11 cm, respectively). For both the DL-S and DL-OBS groups no lung reduction surgery was performed, and in all cases the donor's lungs closely matched the recipient's chest cavity.

Pulmonary Function

Pulmonary function results from the three groups are presented in Table 2. These data are expressed as percentage of predicted of the recipient and presented as mean ± SD values, with comparisons between each group and control subjects. Of the clinically stable group (DL-S), six subjects had normal PFT and as a group many of the lung volumes and spirometric values were similar to control measurements. However, there was evidence of mild pulmonary restriction and gas trapping, as evidenced by mildly reduced TLC and inspiratory capacity (IC) and slow vital capacity (SVC) values, and increased ratio of residual volume to total lung volume (RV/TLV) and decreased FRC_He/FRC_BP ratio, respectively.The N₂SP3 was significantly increased in the DL-S subjects.

In contrast to the above observations in the DL-S group, the patients with obliterative bronchiolitis syndrome (DL-OBS group) demonstrated profound airways obstruction with significantly reduced FEV₁, FEV₁/FVC ratio, forced expiratory flow at 25 to 75% of vital capacity (FEF _25-75%), and sGAW (Table 2). Moreover, the DL-OBS group had markedly more gas trapping, given by an enhanced RV/TLC ratio and decreased FRC_He/FRC_BP. They also had markedly increased N₂SP3 at more than 7-fold above control subjects. Both resting Sa_O2 and PET_CO2 measurements were within normal range in the DL-OBS patients, however, these were significantly lower than values in control subjects.

In light of the above observations, we examined all data from the DL-S and DL-OBS subjects collectively to assess for the presence of small airways dysfunction. A N₂SP3 value of 1.5% or greater derived from our control group (mean ± 2SD) and similar to other studies (15, 16) was used as a cutoff point indicating small airways dysfunction. This analysis showed that 13 of 15 (87%) of all DL patients, 9 of 11 (82%) patients of the DL-S group, and 4 of 4 (100%) patients of the DL-OBS group had significantly elevated N₂SP3. Moreover, 4 of the 6 patients in the DL-S group with normal PFT (percentage of predicted > 80%) had abnormally elevated N₂SP3. In addition, the correlation between N₂SP3 and various PFT measurements was calculated for all patients collectively: FEV₁ percentage of predicted (r = 0.58; p = 0.024; Figure 2A); FEV₁/FVC (r = 0.63; p = 0.011; Figure 2B); RV/TLC (r = 0.72; p = 0.002; Figure 2C).

View larger version (26K): [in this window] [in a new window]   Figure 2.   Scatter plot of individual slopes of N2SP3 versus corresponding individual FEV1 (A); FEF1/FVC (B); RV/TLC (C ); and FEF25-75% (D) in 15 control subjects (closed circles), 11 DL-S (open triangles), and 4 DL-OBS (closed triangles). Correlation coefficient (r) and 95% confidence intervals for the combined 15 DL-S and DL-OBS subjects are shown.

Pulmonary Mechanics

As indicated in Table 3, static lung compliance determinants of our control subjects were in agreement with those reported in the literature (24). However, the various determinants were significantly reduced in both the DL-S and DL-OBS patients, noting that all these measurements were more profoundly decreased in the DL-OBS group. The exponential slope constant, K, was significantly reduced in the DL-S patients, and was further decreased in the DL-OBS group. Lung recoil pressures at 90% TLC were found to be within normal range established by the control group in all subjects studied except for one in the DL-OBS group which reduced significantly the mean recoil pressure of this small group. Dynamic compliance (Cdyn,L) at tidal volume was significantly reduced in both the DL-S and DL-OBS groups. However, the ratio of Cdyn,L/Cst,L at tidal breathing did not differ from control subjects.

Upstream pulmonary resistance (R_US) was markedly increased in the DL-OB group, whereas R_US was not significantly altered in the DL-S group. However, we found a significant positive correlation for all transplant subjects between R_US and N₂SP3 (r = 0.61; p = 0.016).

Finally, we found significant correlation for all DL-S and DL-OB subjects between FEV₁ (percentage of predicted) and various compliance measurements: Cst,L (r = 0.61; p = 0.016; Figure 3A); SCst,L (r = 0.55; p = 0.032; Figure 3B), Cst,L/ TLC (r = 0.53; p = 0.04; Figure 3C).

View larger version (28K): [in this window] [in a new window]   Figure 3.   Scatter plot of individual slopes of FEV1 versus corresponding individual Cst,L (A); SCst,L (B); Cst,L/TLC (C ) in 15 control subjects (closed circles), 11 DL-S (open triangles), and four DL-OBS (closed triangles). Correlation coefficient (r) and 95% confidence intervals for the combined 15 DL-S and DL-OBS subjects are shown.

	DISCUSSION

TOP ABSTRACT INTRODUCTION METHODS RESULTS DISCUSSION REFERENCES

This is the first comprehensive study to evaluate lung mechanics in stable patients after DL transplantation. We found reduced compliance measurements near FRC, with normal elastic recoil pressures at high lung volumes. In addition, based on the single-breath nitrogen test we have determined significant small airways dysfunction in the majority of patients.

All patients, including the DL-OBS group, were exceptional in their relative good health. They were 2 ¹/₂ to 3 yr post-transplant and had no symptoms of acute rejection or respiratory infections in the previous year. Six subjects in the DL-S group had normal PFT and as a group the DL-S subjects had only mild pulmonary restriction compared with control subjects. In contrast, significant obstruction lung disease was noted in all DL-OBS subjects.

It is known that with time, 20 to 40% of all patients after lung transplantation will advance to OB (4) with deterioration in pulmonary status (4). Early recognition of pre-OB stages is essential, although, at present, we are limited in the treatment of OB. Moreover, it seems that by the time the diagnosis is established the process has progressed to an irreversible stage. Early studies have attempted to determine early signs of OB. Transbronchial biopsy is the "gold standard," however, this procedure is invasive, has significant sampling error, and requires experienced centers to perform and grade the results (6, 9). Therefore, spirometric criteria have been established for the diagnosis and staging of OBS (3). A drop in 20% from the two best post-transplant values of FEV₁ was chosen by consensus as indicating OBS. A recent study on stable lung transplant recipients suggests that a drop of 12% in FEV₁ from post-transplant baseline values is indicative of a significant decrease in allograft function that may be due to infection or rejection (28). In addition, FEF_25-75% and sGAW have been suggested to detect early allograft dysfunction (29, 30).

Obstructive lung disease is the clinical manifestation of patients with OB or OBS. Therefore, we wondered if our subjects who were relatively stable 2 to 3 yr post-transplant and especially those with normal PFT had evidence of small airway dysfunction reflected by the single-breath nitrogen test. Using this technique, we determined that 82% of the DL-S subjects and 100% of the DL-OBS subjects had small airway dysfunction. Moreover, 4 of the 6 patients in the DL-S group with normal PFT had abnormal N₂SP3. Additional support for small airways dysfunction in our patients came from our findings of a positive correlation between R_US which reflects peripheral airway patency (23) and N₂SP3, and significant obstructive lung disease seen in the DL-OBS group and some of the DL-S patients based upon interpretation of their PFT. A question to consider is whether the overall differences between the groups' pulmonary function represent different clinical entities, or if small airways dysfunction found in both groups represents a continuum of pathology.

Pulmonary function has been studied after DL transplantation. Mild restrictive lung disease was reported in seven stable patients who were followed for 6 mo after transplant (31). Two mechanisms were proposed to explain these early restrictive findings. The first involves altered chest wall mechanics post-thoracotomy similar to those described in patients after open heart surgery. The second mechanism is related to disparity between the volume of the donor's lungs and the volume of the thoracic cavity of the recipient. In contrast to the mild restrictive findings in stable patients, patients with OB followed for 8 to 36 mo showed severe obstructive lung disease similar to the findings in the DL-OBS group (32).

The correlations between N₂SP3 as an indicator of small airways dysfunction and pulmonary function parameters suggest that small airways function in both our groups may follow a progressive pattern of deterioration. The regression intercepts in Figures 2A-C suggest that patients with no evidence of small airways dysfunction per N₂SP3 value would have normal PFT values, whereas DL-S and DL-OBS subjects exhibit gradual alterations in pulmonary function parameters suggesting progressive obstructive lung disease. The pattern in Figure 2D suggests that early small airways dysfunction measured by N₂SP3 correlates with an early fall in FEF_25-75%, as has been suggested by others as an early indicator for the development of OB (29).

Our findings of reduced compliance measurement near FRC for both the DL-S and DL-OBS groups and normal elastic recoil pressures at 90% TLC in 14 of 15 subjects do not suggest intrinsic changes in lung parenchyma. This was also apparent from the shape and position of the pressure-volume curves for these patients, which do not follow a pattern for restrictive lung disease. Several factors may have contributed to reduced compliance in our study. Lungs from younger donors could account for some of the decreased compliance. Analysis based on a standard compliance/age regression shows that for our study, donor youth may account for about 4% and 6% of the reduction in the DL-S and DL-OBS measurements whereas we found 29% and 53% reduction in these groups, respectively. Another possibility is the fact that both groups had evidence of air trapping and hyperinflation; this could have reduced chord compliance measurements by an upward shift of the compliance curve. However, this would not affect SCst,L and Cst,L/TLC which are normalized for volumetric changes.

In our opinion, structural or functional abnormalities in the small airways rather than intrinsic changes in the lung parenchyma distal to these airways are the main factor to account for reduced compliance in our patients. Several models support this speculation, linking the contribution of small airways to mechanical properties of the lung. The distal airways of the tracheobronchial tree make a small but significant contribution to lung compliance, especially at the low lung volumes where chord compliance was measured (18). Under conditions affecting the integrity or function of these airways, elastic recoil properties may be altered by more than 20% mainly by deforming the lung parenchyma (33, 34).

We are aware that our data were obtained as a cross-section, and a longitudinal study is indicated in order to assess for progressive lung dysfunction after DL transplantation. However, given this limitation, the distribution of small airways dysfunction and compliance determinants between and within the DL-S and DL-OBS groups suggest a similar pattern of progressive lung dysfunction after DL transplantation.

In summary, we have demonstrated significant small airways dysfunction and significant alterations in lung mechanics in patients after DL transplantation. We speculate that reduced compliance measurements found in these patients are linked to alterations in the small airways. Of special concern are our findings that the DL-S group had significant changes in compliance and small airways function in a pattern similar to the DL-OB group, only smaller in magnitude.