The mechanisms driving the transition from inflammation to fibrosis in sarcoidosis patients are poorly understood; prognostic features are lacking. Immune cell profiling may provide insights into pathogenesis and prognostic factors of the disease. This study aimed to establish associations in simultaneous of lymphocyte subset profiles in the blood, bronchoalveolar lavage fluid (BALF), and lung biopsy tissue in the patients with newly diagnosed sarcoidosis.A total of 71 sarcoid patients (SPs) and 20 healthy controls (HCs) were enrolled into the study. CD31, CD38, CD44, CD103 positive T lymphocytes in blood and BALF were analysed. Additionally, the densities of CD4, CD8, CD38, CD44, CD103 positive cells in lung tissue biopsies were estimated by digital image analysis.Main findings: (I) increase of percentage of CD3+CD4+CD38+ in BALF and blood, and increase of percentage of CD3+CD4+CD44+ in BALF in Löfgren syndrome patients comparing with patients without Löfgren syndrome, (II) increase of percentage of CD3+CD4+103+ in BALF and in blood in patients without Löfgren syndrome (comparing with Löfgren syndrome patients) and increase of percentage of CD3+CD4+103+ in BALF and in blood in more advanced sarcoidosis stage. (III) Increasing percentage of BALF CD3+CD4+CD31+ in sarcoidosis patients when comparing with controls independently of presence of Löfgren syndrome, smoking status or stage of sarcoidosis. Several significant correlations were found.Lymphocyte subpopulations in blood, BALF, and lung tissue were substantially different in SPs at the time of diagnosis compared to HCs. CD3+CD4+CD31+ in BALF might be a potential supporting marker for the diagnosis of sarcoidosis. CD3+CD4+CD38+ in BALF and blood and CD3+CD4+CD44+ in BALF may be markers of the acute immune response in sarcoidosis patients. CD4+CD103+ T-cells in BALF and in blood are markers of the persistent immune response in sarcoidosis patients and are potential prognostic features of the chronic course of this disease.
The diagnosis of lung sarcoidosis relies in part on the observation of alveolar CD4+ cells and the demonstration of an increased CD4/CD8 ratio. This ratio has been proposed as a diagnostic tool for pulmonary sarcoidosis. But this anomaly is also found in other lung diseases too. The search for other pathognomonic criteria allowing the discrimination of sarcoidosis patients (pts.) from patients with CD4+ alveolar lymphocytosis has been disappointing. We investigated CD103 molecules on the T lymphocytes subpopulations. The expression of these molecules was examined on BAL lymphocytes from sarcoid patients with different radiologycal stages (Ist. – 23 pts; II – 16 pts: III – 9 pts.) and Lefgren9s sindrome (25 pts) and patients with other lung disemination (17 pts.). For all patients, the expression of CD3, CD4, CD8 and CD103 was assessed by flow cytometry. We found that CD4+/CD8+ ratio in I stage was 6,5±4,1; II st. – 4,5±2,3, III st. – 4,1±3,3, Lefgren9s sindrome – 8,6±4,9 and other lung disemination – 1,9±1,7. CD4+CD103+/CD4+ ratio in I st. – 0,2±0,1; 0,2±0,1; 0,2±0,1; 0,1±0,1 and 0,4±0,2 respectively. CD4+CD103+/CD4+ ratio differ significally (p Our findings demonstrate that the combined use of CD4+/CD8+ and CD4+CD103+/CD4+ ratios provides a highly sensitive indicator of lung sarcoidosis in patients with other CD4+ BAL lymphocytosis.
Reikšminiai žodžiai: anti-ŽLA antikūnai, inksto transplantato atmetimas, Luminex technologija. Įvadas. Ūminis inksto transplantato atmetimas diagnozuojamas remiantis Banff kriterijais, į kuriuos įtraukti donorui specifiniai antikūnai ir histologiniai radiniai. Antikūnai prieš žmogaus leukocitų antigenus (anti-ŽLA antikūnai), tarp jų ir donorui specifiniai antikūnai, nustatomi Luminex technologija, kuri Vilniaus universiteto ligoninės Santariškių klinikose (VUL SK) pradėta taikyti 2012 m. Mūsų atlikto tyrimo tikslas yra įvertinti anti-ŽLA antikūnų ryšį su histologiniais radiniais bei paanalizuoti gydymo taktikos parinkimą atsižvelgiant į imunologinio ir histologinio tyrimo rezultatus pacientams po inksto transplantacijos. Tyrimo medžiaga ir metodai. Atliktas retrospektyvinis tyrimas VUL SK. Tiriamųjų grupę sudarė 65 pacientai, kuriems 2014– 2015 m. atlikta inksto transplantato biopsija bei imunologinis tyrimas (Luminex, kryžminės dermės mėginys limfocitotoksiniu arba tėkmės citometrijos metodu) biopsijos metu. Į tyrimą pagal šiuos kriterijus įtrauktos 89 biopsijos. Rezultatai. Iš 89 histologinių tyrimų gauti atsakymai: norma – 1 (1,1 proc.), transplantato atmetimas – 39 (43,8 proc.), lėtinė nefropatija/glomerulopatija – 20 (22,5 proc), kita – 29 (32,6 proc.). Iš visų imunologinių tyrimų 46 buvo atlikti atrankine Luminex techologija (iš jų 32 (69,6 proc.) buvo teigiami), 24 – specifiškumo Luminex technologija (iš jų 21 (87,5 proc.) buvo teigiamas). Luminex teigiamoje grupėje histologiniai rezultatai pasiskirstė taip: ptc0 – 65 proc., ptc1 – 29,3 proc., ptc2 – 4,9 proc. (Luminex neigiamoje – ptc0 – 76,9 proc., ptc1 – 23,1 proc., ptc2 – 0 proc., p > 0,05); g0 – 61 proc., g1 – 29,3 proc., g2 – 7,3 proc., g3 – 2,4 proc. (Luminex neigiamoje grupėje – g0 – 84,6 proc., g1 – 15,4 proc., g2 and g3 – 0 proc., p > 0,05). Esant žymiam C4d teigiamumui (> 51 proc.), visais atvejais tiriant Luminex technologija gautas teigiamas atsakymas. GFG biopsijos metu nuo bazinio GFG Luminex teigiamoje grupėje vidutiniškai sumažėjo 2,8 ± 10,3 ml/min/1,73 m2, o Luminex neigiamoje grupėje vidutiniškai padidėjo 2,0 ± 12,2 ml/min/1,73 m2. GFG 3 mėn. po biopsijos Luminex teigiamoje grupėje vidutiniškai padidėjo 10,6 ± 24,3 ml/min/1,73 m2, o Luminex neigiamoje grupėje vidutiniškai padidėjo 6,8 ± 24,3 ml/min/1,73 m2 (p > 0,05). Papildomas imunosupresinis gydymas Luminex teigiamoje ir Luminex neigiamoje grupėse skirtas atitinkamai 53,7 proc. ir 38,5 proc. pacientų (p = 0,263), taip pat 92,1 proc. pacientų, kuriems histologiškai rastas atmetimas, ir 28 proc., kuriems atmetimo nerasta (p = 0,000). Išvados. Nors ir negauta statistiškai reikšmingo skirtumo, pastebima tendencija, jog Luminex teigiamoje grupėje dažniau nustatytas glomerulitas, peritubulinis kapiliaritas bei C4d teigiamumas. Papildomas imunosupresinis gydymas pacientams skirtas labiau atsižvelgiant į histologinius radinius nei į imunologinio tyrimo rezultatus.
Abstract Introduction: Bronchoalveolar lavage (BAL) as a method of sampling cells is useful in the diagnosis and differential diagnosis of sarcoidosis. However, CD4/CD8 ratio in BAL fluid (BALF) is highly variable and it generates continuous discussions about its diagnostic role. Objective: To prospectively evaluate diagnostic role of BALF CD4/CD8 ratio in pulmonary sarcoidosis manifested in different radiographic and clinical forms in the real clinical practice. Material and methods: The study population consisted of 318 sarcoid patients with a newly diagnosed disease. Comparator groups consisted of 55 healthy subjects and 130 patients with other disorders who underwent BAL and examination of CD4/CD8 ratio in BALF as a step of diagnostic pathway. Diagnostic accuracy of CD4/CD8 ratio in BALF using receiver‐operating characteristic analysis has been calculated. Results: The percentage of BALF lymphocytes in sarcoid patients was significantly different from comparator groups. Normal BALF cell counts were found in 7% of sarcoid patients. However, typical sarcoid BALF cellular pattern was found in 6.2% of all control subjects. We have found that optimal cutoff points for CD4/CD8 ratio are 3.5 and 4.0 for asymptomatic and symptomatic patients, respectively. Sensitivity of the optimal cutoff points of CD4/CD8 ratio was lower in asymptomatic patients compared with symptomatic patients. Sensitivity of the optimal cutoff points decreased with the increased stage of sarcoidosis. Conclusions: BAL is a valuable method in diagnostic pathway of pulmonary sarcoidosis. However, results of BALF examination must be interpreted considering a specific clinical case. BALF CD4/CD8 ratio depends on clinical and radiographic manifestation. Please cite this paper as: Danila E, Norkūnienė J, Jurgauskienė L and Malickaitė R. Diagnostic role of BAL fluid CD4/CD8 ratio in different radiographic and clinical forms of pulmonary sarcoidosis. The Clinical Respiratory Journal 2009; 3: 214–221.
Background. Sarcoidosis is associated with an increase in the number of alveolar T cells (CD3+ cells) and an increase of the CD3+CD4+ lymphocyte subset. However, the number of lymphocytes and the CD4/CD8 ratio in bronchoalveolar lavage (BAL) fluid are highly variable in sarcoidosis. Comparative studies have demonstrated that geographic and ethnic factors are linked to the specific characteristics of patients with sarcoidosis. Aim of the study. To investigate peculiarities of BAL fluid (BALF) cell patterns in different clinical activity of pulmonary sarcoidosis at the time of diagnosis. Material and methods. A total of 308 non-treated patients (138 asymptomatic and 170 with sarcoidosis-related symptoms) and 40 previously empirically steroid-treated patients with newly diagnosed sarcoidosis have been prospectively examined. Results. Significant BAL fluid lymphocytosis and increased CD4/CD8 ratio were characteristic for all three sarcoidosis patient groups. A total of 12% of asymptomatic patients, 3% of patients with sarcoidosis-related symptoms, and 5% of previously treated symptomatic patients had normal BALF cell counts. Non-treated patients with sarcoidosis-related symptoms had significantly higher lymphocytosis (45±19% versus 39±17%, P<0.01), CD4/CD8 ratio (9.3±5.0 versus 5.7±4.5, P<0.001), and total BALF cell count (411±322 106/mL versus 334±273 106/mL, P<0.05), compared with asymptomatic patients. However, previously treated symptomatic patients had lower lymphocytosis (39±15% versus 45±19%, P=0.058), and total BALF cell count (292±166 106/mL versus 411±322 106/mL, P<0.05) compared with non-treated symptomatic patients. The same trend was noticed for CD4/CD8 ratio (8.3±4.8), although a statistically significant difference was not achieved. Conclusions. Independently of clinical symptoms at the time of diagnosis sarcoid patients have significantly different BAL fluid cell patterns compared to healthy persons. BAL fluid cell changes are more prominent in corticosteroid non-treated patients with clinically active sarcoidosis. Treatment with systemic corticosteroids may modify typical BALF cellular patterns of sarcoidosis.
Background: The aim of the present study was to identify specious radiologic and/or physiologic prognostic marker(s), which lead to optimize of the patient follow-up frequency. Methods: Eighty consecutive patients with newly diagnosed pulmonary sarcoidosis. Patients underwent chest radiography, high-resolution computed tomography (HRCT) examination, pulmonary function tests (PFT), bronchoscopy with bronchoalveolar lavage (BAL) and lung biopsy, and bronchoalveolar lavage fluid (BALF) cell examination. Results: The reduction in PFT values seen in radiological sarcoidosis stage III was greater than that seen in stages I and II. The percentage of neutrophils in the lungs was found to increase in stages II and III. PFT indices were correlated negatively with the consolidation and ground glass opacities CT scores, but not with the micronodule or macronodule scores. The rise in the percentage of BALF lymphocytes was associated with the restriction pattern of PFT. The diagnostic value of BALF for sarcoidosis was higher when the typical radiologic patterns of stage I disease were found and that smoking decreased the diagnostic value of CD4/CD8 ratio. Conclusions: This study supports the opinion that the staging of the pulmonary sarcoidosis with chest X-rays is still valuable from the prognostic point of view, because significant correlations between the radiologic stages of sarcoidosis and PFT parameters were found. Chest HRCT was significantly superior to chest X-ray in detecting mediastinal and pulmonary parenchymal changes. However, the prognostic role of HRCT needs to be better investigated evaluating serial examinations. Only consolidation and ground glass scores (neither of which are frequently found in sarcoidosis) hold prognostic value, since these were negatively correlated with PFT parameters.
Background: Many theories have been proposed to explain pathogenesis of COPD; however, remains unclear why the majority of smokers (~80 %) do not develop COPD, or only develop a mild disease. To explore if COPD has an autoimmune component, the role of T regulatory lymphocytes (Tregs) in the lung tissue of COPD patients is of a crucial importance. Methods: Bronchial tissue biopsy samples were prospectively collected from 64 patients (39 COPD and 25 controls – 15 smokers and 10 non-smokers). The patients with COPD were subdivided into mild/moderate (GOLD stage I-II) and severe/very severe (GOLD stage III-IV) groups. Digital image analysis was performed to estimate densities of CD4+CD25+ cell infiltrates in double immunohistochemistry slides of the biopsy samples. Blood samples were collected from 42 patients (23 COPD and 19 controls) and tested for CD3+CD4+CD25+bright lymphocytes by flow cytometry. Results: The number of intraepithelial CD4+CD25+ lymphocytes mm-2 epithelium was significantly lower in the severe/very severe COPD (GOLD III-IV) group as well as in the control non-smokers (NS) group (p<0,0001). Likewise, the absolute number of Treg (CD3+CD4+CD25+bright) cells in the peripheral blood samples was significantly different between the four groups (p=0.032). The lowest quantity of Treg cells was detected in the severe/very severe COPD and healthy non-smokers groups. Conclusions: Our findings suggest that severe COPD is associated with lower levels of Tregs in the blood and bronchial mucosa, while higher Tregs levels in the smokers without COPD indicate potential protective effect of Tregs against developing COPD. Keywords: COPD, autoimmunity, T- regulatory cell, bronchial biopsy, flow cytometry
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