IL-12 and IL-18 are increased and stimulate IFN-gamma production in sarcoid lungs

Sarcoidosis is a systemic chronic granulomatous disease of unknown cause. Recent investigations revealed that the cytokine profile in inflamed lesions of sarcoidosis is Th1 dominant. To obtain better immunopathologic understanding of sarcoidosis, we examined the expression of IL-12 and IL-18 and their roles in IFN-gamma production in pulmonary sarcoidosis. Sarcoid cases had significantly elevated levels of IL-12 (p40 and p70) and IL-18 in bronchoalveolar lavage (BAL) fluids compared with healthy subjects. IL-12 p70 and IL-18 were immunohistochemically expressed in the epithelioid cells and giant cells of sarcoid granulomas. Significant induction of IFN-gamma, IL-12 p70, and IL-18 was observed from sarcoid BAL fluid cells with LPS stimulation, whereas LPS tended to induce only IL-12 p70 in BAL fluid cells from healthy subjects. Sarcoid cases had significantly greater IFN-gamma induction with LPS stimulation than healthy subjects did. IL-18 mRNA expression was observed in freshly isolated sarcoid BAL fluid cells as well as in LPS-stimulated sarcoid BAL fluid cells, but IFN-gamma and IL-12 mRNA expression was observed only in LPS-stimulated BAL fluid cells. Treatment with anti-IL-12- and anti-IL-18-neutralizing Abs significantly inhibited IFN-gamma production from LPS-stimulated BAL fluid cells of sarcoid cases. Coadministration of rIL-12 or rIL-18 induced greater IFN-gamma production in sarcoid BAL fluid cells than in normal BAL fluid cells. We concluded that bioactive IL-12 and IL-18 were produced in sarcoid BAL fluid cells and synergistically induced IFN-gamma production, indicating important cytokines in the Th1 response of sarcoidosis.     

Neurotrophins and neurotrophin receptors in pulmonary sarcoidosis - granulomas as a source of expression

Background

Pulmonary sarcoidosis is an inflammatory disease, characterized by an accumulation of CD4⁺ lymphocytes and the formation of non-caseating epithelioid cell granulomas in the lungs. The disease either resolves spontaneously or develops into a chronic disease with fibrosis. The neurotrophins nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) have been suggested to be important mediators of inflammation and mediate tissue remodelling. In support of this, we have recently reported enhanced NGF levels in the airways of patients with pulmonary sarcoidosis. However, less is known about levels of BDNF and NT-3, and moreover, knowledge in the cellular sources of neurotrophins and the distribution of the corresponding neurotrophin receptors in airway tissue in sarcoidosis is lacking.

Methods

The concentrations of NGF, BDNF and NT-3 in bronchoalveolar lavage fluid (BALF) of 41 patients with newly diagnosed pulmonary sarcoidosis and 27 healthy controls were determined with ELISA. The localization of neurotrophins and neurotrophin receptors were examined by immunohistochemistry on transbronchial lung biopsies from sarcoidosis patients.

Results

The sarcoidosis patients showed significantly enhanced NT-3 and NGF levels in BALF, whereas BDNF was undetectable in both patients and controls. NT-3 levels in BALF were found higher in patients with non-Löfgren sarcoidosis as compared to patients with Löfgren''s syndrome, and in more advanced disease stage. Epithelioid cells and multinucleated giant cells within the sarcoid granulomas showed marked immunoreactivity for NGF, BDNF and NT-3. Also, immunoreactivity for the neurotrophin receptor TrkA, TrkB and TrkC, was found within the granulomas. In addition, alveolar macrophages showed positive immunoreactivity for NGF, BDNF and NT-3 as well as for TrkA, TrkB and TrkC.

Conclusions

This study provides evidence of enhanced neurotrophin levels locally within the airways of patients with sarcoidosis. Findings suggest that sarcoid granuloma cells and alveolar macrophages are possible cellular sources of, as well as targets for, neurotrophins in the airways of these patients.     

Pulmonary hypertension associated with sarcoidosis

Pulmonary involvement is common in sarcoidosis, an immune-mediated inflammatory disorder that is characterized by non-caseating granulomas in tissue. Sarcoid patients with advanced pulmonary disease, especially end-stage pulmonary fibrosis, risk developing pulmonary hypertension (World Health Organization group III pulmonary hypertension secondary to hypoxic lung disease). Increased levels of endothelin (ET)-1 in plasma and bronchoalveolar lavage of some sarcoid patients suggest that ET-1 may be driving pulmonary fibrosis and sarcoidosis-associated pulmonary hypertension. Although a relationship between raised levels of ET-1 and clinical phenotype is yet to be identified, early evidence from studies of ET-1 blockade with drugs such as bosentan is encouraging. Such therapy possibly could be combined with standard anti-inflammatory agents to improve outcome.     

Fine needle aspiration biopsy in the diagnosis of sarcoid of the head and neck

The use of fine needle aspiration biopsy to confirm the clinical diagnosis of six cases of sarcoidosis with palpable disease in the head and neck area is reported. The procedure was simple and inexpensive, and the interpretation of the smears and identification of typical sarcoid granulomas was easy.     

Polymorphisms at position -308 in the promoter region of the TNF-alpha and in the first intron of the TNF-beta genes and spontaneous and lipopolysaccharide-induced TNF-alpha release in sarcoidosis.

TNF-alpha is a potent pro-inflammatory cytokine. Previous studies have proved that biallelic polymorphisms in the TNF-alpha (-308, TNFA) and TNF-beta genes (intron 1, TNFB) influence TNF-alpha production. In sarcoidosis, a chronic granulomatous disease, as a result of an unknown in vivo activation bronchoalveolar lavage (BAL) cells release high amounts of TNF-alpha, spontaneously and after in vitro stimulation. Thus, sarcoidosis could serve as a model to test the in vivo effect of TNF gene polymorphisms. We determined the TNFA and TNFB polymorphisms of 44 patients with sarcoidosis and found the following allele frequencies: 0.80, 0.20, 0.38 and 0.62 for TNFA1, TNFA2, TNFB1 and TNFB2, respectively. To examine the in vivo effect of the named polymorphisms on the TNF-alpha production, the spontaneous and LPS-induced TNF-alpha release of BAL cells and peripheral blood mononuclear cells were also determined in patients with sarcoidosis. Statistical analysis did not reveal any significant difference between sarcoidosis patients with different genotypes. The results show that TNFA and TNFB polymorphisms do not determine the level of TNF-alpha release of mononuclear cells activated during the course of sarcoid inflammation.     

Concomitant association of thyroid sarcoidosis and Graves' disease

OBJECTIVE: Graves' disease (GD) with sarcoid involvement of the thyroid gland has rarely been reported. METHOD: We report a case of GD with thyroid sarcoidosis in a 28-year-old woman. Thyroid function was assessed by triiodothyronine (T(3)), thyroxine (T(4)), thyroid-stimulating hormone (TSH) and TSH receptor antibodies (TSH-R Ab). Thyroid scintigraphy, ultrasound and fine-needle aspiration biopsy were performed. The patient underwent surgery. RESULT: The patient had a nodular goiter. Serum T(3), T(4) and TSH-R Ab levels were elevated with suppressed TSH level. Scintigraphy showed diffuse activity as seen in GD, and ultrasound revealed that parenchyma was heterogenous. Sarcoidosis was discovered on routine chest X-ray. Although no sarcoid involvement was found on specimen, the thyroid gland showed non-caseating granulomas on histology. CONCLUSION: Since sarcoid involvement of the thyroid gland can cause hypofunction, we report the uncommon infiltration of sarcoidosis with hyperthyroidism.     

Mycobacterium tuberculosis DNA in tissue affected by sarcoidosis.

OBJECTIVE--To investigate the prevalence of Mycobacterium tuberculosis DNA in granulomatous tissues from patients with sarcoidosis and from controls matched for age, sex, and tissue by using the polymerase chain reaction. DESIGN--Single blind control trial. SUBJECTS--16 patients with sarcoidosis who had undergone diagnostic biopsy of lung, skin, or lymph node and 16 patients with squamous cell carcinoma or Hodgkin''s disease to act as controls. In addition, four lung specimens infected with M tuberculosis were included as positive controls. RESULTS--M tuberculosis DNA was present in sarcoid tissues containing granulomas from seven of the 16 patients and one of the 16 matched controls. Two of the four specimens known to be infected with M tuberculosis were positive in the controlled experiment. CONCLUSION--These figures suggest that M tuberculosis DNA is detected as readily in patients with sarcoidosis as in patients with frankly tuberculous tissues and imply that M tuberculosis may be linked to the cause of sarcoidosis.     

Unusual Clinical Course of Graves’ Thyrotoxicosis and Concomitant Sarcoidosis: Case Report and Review of Literature

ObjectiveTo report a case of Graves’ disease with concomitant sarcoidosis involving the thyroid gland.MethodsWe present the clinical, laboratory, imaging, and pathologic findings and describe the clinical course of a patient with Graves’ disease and sarcoidosis, who was unresponsive to propylthiouracil and radioiodine treatment.ResultsA 23-year-old woman presented with thyrotoxicosis and a large goiter. Laboratory studies and findings on thyroid uptake and scan were consistent with Graves’ disease. She was also found to have hilar lymph-adenopathy and hepatosplenomegaly. Despite treatment with antithyroid drugs and radioiodine therapy, her hyperthyroidism persisted. Surgical resection of the thyroid gland and 2 lymph nodes disclosed noncaseating granulomas, consistent with sarcoid.ConclusionAutoimmune endocrinopathies and, less commonly, thyroid autoimmune disease have been reported in patients with sarcoidosis. Similarities exist in the pathogenesis of these two conditions. Concomitant sarcoidosis in the thyroid gland in patients with Graves’ disease may contribute to the resistance to antithyroid drugs and radioiodine therapy. (Endocr Pract. 2007;13:159-163)     

Intracellular cytokine profiles and T cell activation in pulmonary sarcoidosis

In granulomatous inflammatory lung diseases such as sarcoidosis, the balance of cytokine production by activated T cells in the lungs may influence clinical disease outcome. To investigate the potential of T lymphocytes to produce cytokines and contribute to this process, T cells from bronchoalveolar lavage (BAL) and PB from 19 patients with active lung disease were stimulated, stained, and analysed by flow cytometry for intracellular production of cytokines and expression of the activation marker CD69. Higher proportions of BAL cells expressed CD69 compared with PB, in the absence of in vitro stimulation. The expression of IFN-γ was similar in unstimulated BAL and PB T cells, and there was no association between the expression of CD69 and IFN-γ. Following stimulation, there were increased numbers of IFN-γ⁺ T cells. A similar trend was found with IL-2⁺ T cells, but there were lower levels of IL-4⁺ T cells in BAL compared with PB, and similar levels of IL-10⁺ T cells. The presence of activated T lymphocytes in BAL samples from patients with sarcoidosis, with the potential to produce Th1 type 1 cytokines may contribute to the inflammatory processes in this granulomatous lung disease. The use of intracellular flow cytometry to investigate cytokine production by BAL T cells could help to indicate potential targets for future therapy.     

Leukocyte immunophenotypes in bronchoalveolar lavage fluid and peripheral blood of paracoccidioidomycosis, sarcoidosis and silicosis.

Leukocyte subsets in bronchoalveolar lavage (BAL) fluid and peripheral blood of patients with paraccoccidioidomycosis, sarcoidosis and silicosis were characterized using monoclonal antibodies and an immunoperoxidase technique. In paraccocidioidomycosis, the number of T-helper/inducer CD4-positive lymphocytes was lower in peripheral blood than in BAL fluid. Additional analysis showed that the expression of HLA-DR was very similar in alveolar macrophages, lung and blood T-cells. In sarcoidosis and silicosis there were higher proportions of T-helper/inducer cells in peripheral blood than in BAL fluid. The alterations in the T-helper/inducer/T-suppressor/cytoxic CD4/CD8 ratio in sarcoidosis and silicosis were more appreciable in peripheral blood than in BAL fluid, contrasting with the results in paracoccidioidomycosis. The expression of HLA-DR by alveolar macrophages in sarcoidosis was the highest of all the disease studied. No statistically significant differences were observed between chronic multifocal and chronic unifocal paracoccidioidomycosis disease, stage II and stage III sarcoidosis, and chronic and accelerated silicosis. The three granulomatous diseases analyzed had a few alveolar macrophages expressing the CD4 molecule on their surface. These findings and the technique of analyzing both peripheral blood and BAL leukocyte subsets may help to understand the pathogenesis of interstitial lung diseases.     

In vitro culture of leukocytes from patients with sarcoidosis: in search of an "in vitro Kveim test"

Mononuclear cells from patients with sarcoidosis respond to Kveim tissue suspensions in vivo and from granulomas. We attempted to develop an in vitro Kveim test by adding Kveim material to monocytes and lymphocytes from patients with sarcoidosis and observing the cells in tissue culture. Sarcoid leukocytes were grown in culture with either Kveim suspensions (100 microgram/ml), control human spleen extract (100 microgram/ml) or no foreign antigen. After 7 to 9 days, we counted the number of cells adherent to glass, the number of cells with nucleoli, and the number of giant cells. Despite equal monocyte inocula, cultures of sarcoid leukocytes consistently had more cells adherent to glass than did cultures from normal donors regardless of whether Kveim, spleen, or no antigen was added to the cultures. There were no significant differences in giant cell formation or in the number of cells with nucleoli in the sarcoid compared with the control cultures.     

Sarcoidosis in Native and Transplanted Kidneys: Incidence,Pathologic Findings,and Clinical Course

Renal involvement by sarcoidosis in native and transplanted kidneys classically presents as non caseating granulomatous interstitial nephritis. However, the incidence of sarcoidosis in native and transplant kidney biopsies, its frequency as a cause of end stage renal disease and its recurrence in renal allograft are not well defined, which prompted this study. The electronic medical records and the pathology findings in native and transplant kidney biopsies reviewed at the Johns Hopkins Hospital from 1/1/2000 to 6/30/2011 were searched. A total of 51 patients with a diagnosis of sarcoidosis and renal abnormalities requiring a native kidney biopsy were identified. Granulomatous interstitial nephritis, consistent with renal sarcoidosis was identified in kidney biopsies from 19 of these subjects (37%). This is equivalent to a frequency of 0.18% of this diagnosis in a total of 10,023 biopsies from native kidney reviewed at our institution. Follow-up information was available in 10 patients with biopsy-proven renal sarcoidosis: 6 responded to treatment with prednisone, one progressed to end stage renal disease. Renal sarcoidosis was the primary cause of end stage renal disease in only 2 out of 2,331 transplants performed. Only one biopsy-proven recurrence of sarcoidosis granulomatous interstitial nephritis was identified.

Conclusions

Renal involvement by sarcoidosis in the form of granulomatous interstitial nephritis was a rare finding in biopsies from native kidneys reviewed at our center, and was found to be a rare cause of end stage renal disease. However, our observations indicate that recurrence of sarcoid granulomatous inflammation may occur in the transplanted kidney of patients with sarcoidosis as the original kidney disease.     

The anergic state in sarcoidosis is associated with diminished dendritic cell function

Sarcoidosis is a chronic inflammatory disease of unknown cause, characterized by granuloma formation similar to tuberculosis, but without clear evidence of a microbial infection. Because sarcoidosis is linked with clinical anergy and other evidence of diminished cellular immunity, we hypothesized that decreased skin delayed-type hypersensitivity (DTH) responses to recall Ags in affected individuals would be associated with decreased function of their blood dendritic cells (DCs). Our study involved ex vivo isolation, phenotyping, and functional testing of myeloid DCs (mDCs), plasmacytoid DCs, and T lymphocytes from blood of normal healthy volunteers and sarcoidosis subjects with active, untreated pulmonary disease. We found mDC function in the allogeneic MLR directly corresponded to the magnitude of skin DTH reactions to recall Ags in both sarcoidosis subjects and normal volunteers. However, both of these outcomes were significantly decreased in the sarcoidosis group. Diminished mDC function occurred despite up-regulated costimulatory and maturation markers. Clinical relevance is suggested by the inverse relationship between both mDC allogeneic responses and skin DTH responses with clinical disease severity as measured by chest radiograms. Because granulomas form when cellular immunity fails to clear antigenic stimuli, attenuated mDC function in sarcoidosis may contribute to susceptibility and persistence of the chronic inflammation characteristic of this disease.     

Bronchoalveolar cells from sarcoid patients demonstrate enhanced antigen presentation

The recognition of foreign antigens by T lymphocytes in association with lung antigen-presenting cells may be critical in the initiation of the mononuclear alveolitis and granuloma formation of pulmonary sarcoidosis. However, it has been shown that bronchoalveolar cells (BAC) from normal volunteers function poorly as antigen-presenting cells. Therefore, the ability of sarcoid BAC to serve as accessory cells for antigen-dependent autologous T cell proliferation, as measured by tritiated thymidine uptake, was compared with that of normal BAC. Although irradiated sarcoid BAC supported antigen-induced T cell proliferation, normal BAC did so poorly (p less than 0.005). Because it has been shown that sarcoid BAC produce more interleukin 1 (IL 1) than normal BAC, it was considered that the enhancement of antigen-induced proliferative responses could result from an increased amount of IL 1, and that contaminating monocytes in the peripheral blood T cell preparations displayed the antigen for T cell recognition. Therefore, it was necessary to establish that antigen-induced T cell responses required HLA-D region compatibility between the sarcoid BAC and T lymphocytes. BAC from sarcoid patients stimulated antigen-specific proliferation in T cells lines matched for at least one HLA-D-region antigen, but failed to stimulate T cell lines that were unmatched for both antigens. This finding indicates that cells in bronchoalveolar lavage fluids from sarcoid patients were fully capable of acting as antigen-presenting cells. The identification of antigen-presenting cells in the lungs of patients with sarcoidosis together with the previous findings of activated T cells, enhanced IL 1 production, and spontaneous interleukin 2 release in sarcoid patients is compatible with the hypothesis that local cell-mediated immunity is involved in the pathogenesis of pulmonary sarcoidosis.     

Reactive type II pneumocytes in bronchoalveolar lavage fluid

OBJECTIVE: To evaluate the prevalence of reactive type II pneumocytes (RPII) in bronchoalveolar lavage (BAL) fluid samples obtained from patients with various pulmonary disorders. STUDY DESIGN: Consecutive BAL fluid samples were screened for the presence of RPII on May-Grünwald-Giemsa-stained cytocentrifuge preparations. BAL fluid samples with and without RPII were compared with regard to prevalence, associated clinical diagnoses and cytologic findings. RESULTS: RPII were generally large cells with a high nuclear:cytoplasmic ratio and deeply blue-stained, vacuolated cytoplasm. Most RPII occurred in cohesive cell groups, and the vacuoles tended to be confluent. Cytologic findings associated with RPII were foamy alveolar macrophages, activated lymphocytes and plasma cells. RPII were present in 94 (21.7%) of 433 included BAL fluid samples. The highest prevalences were noted in patients with systemic inflammatory response syndrome and alveolar hemorrhage. In addition, RPII tended to occur more frequently in ventilator-associated pneumonia, Pneumocystis carinii pneumonia, extrinsic allergic alveolitis and drug-induced pulmonary disorders. In contrast, RPII were not observed in BAL fluid samples obtained from patients with sarcoidosis. CONCLUSION: RPII were prevalent in about 20% of BAL fluid specimens. They were associated mainly with conditions of acute lung injury and not observed in sarcoidosis.     

Characterization of mononuclear phagocyte subpopulations in the human lung by using monoclonal antibodies: changes in alveolar macrophage phenotype associated with pulmonary sarcoidosis

Current concepts of pulmonary sarcoidosis suggest that the alveolar macrophage plays a central role in the pathogenesis of the disease. To help define the population of alveolar macrophages in sarcoidosis, we compared the surface phenotype of alveolar macrophages from patients with sarcoidosis and from normal individuals by using monoclonal antibodies (63D3, OKM1, M phi P-9, M phi S-1, 61D3, and M phi S-39) that detect surface antigens on cells of monocyte/macrophage lineage. Although almost all blood monocytes expressed surface antigens detected by each of these antibodies, only a minority of normal alveolar macrophages expressed the same surface antigens (p less than 0.05, each comparison). However, in sarcoidosis, the percentage of alveolar macrophages expressing these surface antigens was increased (p less than 0.05, each comparison with normal alveolar macrophages). Several findings supported the conclusion that the increased expression of these monocyte-lineage surface antigens on sarcoid alveolar macrophages resulted from increased recruitment of monocytes to the lung in sarcoidosis and not from abnormal "activation" of alveolar macrophages. First, alveolar macrophages expressing these antigens had an immature morphology. Second, in vitro cultivation of blood monocytes and alveolar macrophages in the presence of immune and inflammatory mediators, including mediators known to be present in the lung in sarcoidosis, did not prevent the loss of expression of monocyte-lineage surface antigens from monocytes or induce reexpression of monocyte-lineage surface antigens on alveolar macrophages. Third, the expression of monocyte-lineage surface antigens was only increased on sarcoid macrophages from patients whose lower respiratory tract contained an increased number of T lymphocytes, cells known to release monocyte chemotactic factor in sarcoidosis. Consistent with the knowledge that corticosteroids usually suppress the alveolitis of active sarcoidosis, when the expression of alveolar macrophage surface antigens was evaluated before and during therapy, the percentage of alveolar macrophages expressing monocyte-lineage surface antigens returned to normal after 1 to 3 mo of therapy.(ABSTRACT TRUNCATED AT 400 WORDS)     

Sarcoidosis and Polyarthritis

Polyarthritis is a well-recognized manifestation of sarcoidosis, but the various series of cases reported in the literature reveal considerable variation in its incidence as well as in its clinical manifestations. Three major types of sarcoid polyarthritis are defined, as distinguished by their clinical and pathological features. Two of these types are usually seen in conjunction with the subacute or transient form of sarcoidosis and, in most cases, are accompanied by erythema nodosum. They differ markedly in severity, but their prognosis is uniformly good. The third type, in contrast, is of a chronic nature and is often associated with permanent joint changes. Mild joint manifestations are a frequent finding in sarcoidosis, but more severe arthritis is relatively rare.Two cases of sarcoidosis, presenting with severe polyarthritis, are reported in detail, and cases seen in Halifax hospitals during the period 1954 to 1964 are reviewed.     

RNA interference as a gene silencing therapy for mutant MYOC protein in primary open angle glaucoma

Bronchoalveolar lavage (BAL) is a useful diagnostic tool in interstitial lunge diseases (ILD). However, differential cell counts are often non specific and immunocytochemistry is time consuming. Staining of glyoproteins by periodic acid Schiff (PAS) reaction may help in discriminating different forms of ILD. In addition, PAS staining is easy to perform. BAL cells from patients with idiopathic pulmonary fibrosis (IPF) (n = 8), sarcoidosis (n = 9), and extrinsic allergic alveolitis (EAA) (n = 2) were investigated. Cytospins from BAL cells were made and cells were stained using Hemacolor quick stain and PAS staining. Lymphocytic alveolitis was found in sarcoidosis and EAA whereas in IPF both lymphocytes and neutrophils were increased. PAS positive cells were significantly decreased in EAA compared to IPF and sarcoidosis (25.5% ± 0.7% vs 59.8% ± 25.1% and 64.0% ± 19.7%, respectively) (P < 0.05). No significant correlation between PAS positive cells and inflammatory cells was observed. These results suggest that PAS staining of BAL cells may provide additional information in the differential diagnosis of ILD. Further studies ware warranted to evaluate PAS staining in larger numbers of BAL from patients with ILD.     

Unusual presentations of inflammatory conditions in cerebrospinal fluid

Unusual inflammatory reactions in cerebrospinal fluid (CSF) in five patients were explicable by the type of intracranial injury or surgical intervention that they had received or by their basic disease process. Lumbar puncture fluid from a 64-year-old man with multiple facial fractures contained neutrophils, bacteria, Candida sp. and ciliated columnar cells, findings consistent with a basilar skull fracture allowing paranasal sinus contents to enter the subarachnoid space. A 59-year-old man with angioimmunoblastic lymphadenopathy developed meningitis and suffered a respiratory arrest; a ventricular fluid contained acute inflammatory cells as well as numerous corpora amylacea. Lumbar CSF obtained during surgery from a 26-year-old man with a pontine glioma contained numerous histiocytes clustered around polarizable filaments, probably strands of gauze introduced during surgery. A specimen of CSF obtained intraoperatively from a 54-year-old man with an acoustic neuroma undergoing a second craniotomy contained multinucleated giant cells bearing suture material. A 19-year-old girl with systemic sarcoidosis had noncaseating granulomas in the right temporal lobe and multinucleated giant cells in her CSF.