Persistence and selection of an expanded B-cell clone in the setting of rituximab therapy for Sjögren's syndrome.

INTRODUCTION: Subjects with primary Sjögren's syndrome (SjS) have an increased risk of developing B-cell lymphoma and may harbor monoclonal B-cell expansions in the peripheral blood. Expanded B-cell clones could be pathogenic, and their persistence could exacerbate disease or predispose toward the development of lymphoma. Therapy with anti-CD20 (rituximab) has the potential to eliminate expanded B-cell clones and thereby potentially ameliorate disease. This study was undertaken to identify and track expanded B-cell clones in the blood of subjects with primary SjS who were treated with rituximab. METHODS: To determine whether circulating B-cell clones in subjects with primary SjS emerge or remain after B cell-depleting therapy with rituximab, we studied the antibody heavy-chain repertoire. We performed single-memory B-cell and plasmablast sorting and antibody heavy-chain sequencing in six rituximab-treated SjS subjects over the course of a 1-year follow-up period. RESULTS: Expanded B-cell clones were identified in four out of the six rituximab-treated SjS subjects, based upon the independent amplification of sequences with identical or highly similar VH, DH, and JH gene segments. We identified one SjS subject with a large expanded B-cell clone that was present prior to therapy and persisted after therapy. Somatic mutations in the clone were numerous but did not increase in frequency over the course of the 1-year follow-up, suggesting that the clone had been present for a long period of time. Intriguingly, a majority of the somatic mutations in the clone were silent, suggesting that the clone was under chronic negative selection. CONCLUSIONS: For some subjects with primary SjS, these data show that (a) expanded B-cell clones are readily identified in the peripheral blood, (b) some clones are not eliminated by rituximab, and (c) persistent clones may be under chronic negative selection or may not be antigen-driven. The analysis of sequence variation among members of an expanded clone may provide a novel means of measuring the chronicity and selection of expanded B-cell populations in humans.

Normative values for exhaled breath condensate pH and its relationship to exhaled nitric oxide in healthy African Americans.

BACKGROUND: Exhaled breath condensate (EBC) pH and exhaled nitric oxide (FeNO) have been proposed as markers of asthma severity. EBC pH values below 6.5 have been associated with asthma exacerbations. Protonation of airway nitrite occurs at low pH and may contribute to FeNO. OBJECTIVE: To establish normative EBC pH values and to determine the contribution of EBC pH to FeNO in healthy African Americans. METHODS: Two hundred seventy healthy African American subjects without asthma between 18 and 40 years old were evaluated. Subjects had simultaneous measurement of EBC pH, EBC nitrite, nitrate, and FeNO. RESULTS: The median EBC pH was 8.14 (interquartile range, 7.83-8.28). Of subjects, 11.9% had an EBC pH < or = 6.5. In subjects with EBC pH values below 6.5, there was an inverse correlation between EBC pH and FeNO (r(2) = 0.158; P = .0245; n = 32). In the entire cohort, there was a direct correlation between EBC pH and EBC nitrite (r(2) = 0.163; P < .0001), but there was no correlation between EBC nitrite and FeNO. In multivariate analyses, EBC pH and nitrite did not contribute significantly to FeNO variation while controlling for other confounders of FeNO. CONCLUSION: There was an increased prevalence (11.9%) of low EBC pH (less than 6.5) in healthy African American subjects compared with white subjects (<5%). EBC pH and nitrite were directly correlated, but there was no correlation between EBC nitrite and FeNO. FeNO correlated with EBC pH at pH values less than 6.5 in univariate but not multivariate analyses. This suggests that EBC pH and nitrite are not significant determinants of FeNO in healthy subjects.

Determinants of exhaled nitric oxide levels in healthy, nonsmoking African American adults.

BACKGROUND: Asthma is a significant cause of morbidity and mortality for African Americans. Fraction of exhaled nitric oxide (FeNO) levels are increased in patients with asthma, and airway levels of nitric oxide metabolites regulate airway inflammation and airway diameter. More needs to be known about the factors that regulate FeNO. There is a need for FeNO reference values for African Americans. OBJECTIVE: We sought to establish reference values and identify factors associated with FeNO levels in healthy African American adults. METHODS: FeNO levels were measured in 895 healthy, nonsmoking African Americans between the ages of 18 and 40 years. FeNO measurements were repeated in 84 subjects. Factors potentially associated with FeNO were measured, including blood pressure, height, weight, and serum total IgE, eosinophil cationic protein, C-reactive protein, and nitrate levels. Data on respiratory symptoms, including upper respiratory tract infection (URI) symptoms, were collected. Univariate and multivariate analyses of the relationship between these variables and FeNO levels were performed. RESULTS: In healthy, nonsmoking African Americans FeNO levels were stable during repeated measurements (intraclass correlation coefficient, 0.81). Sex (P < .0001), serum total IgE levels (P < .0001), and current URI symptoms (P = .0002) contributed significantly to FeNO variability but together accounted for less than 50% of the variation in FeNO levels. CONCLUSION: The high correlation between repeated measurements of FeNO and the low correlation coefficients of known factors associated with FeNO suggest that other factors might contribute substantially to variability of FeNO levels in African Americans.