Expression and functional implications of CCR2 expression on murine alveolar epithelial cells

Acute lung injury results in damage to the alveolar epithelium, leading to leak of proteins into the alveolar space and impaired gas exchange. Lung function can be restored only if the epithelial layer is restored. The process of reepithelialization requires migration of lung epithelial cells to cover denuded basement membranes. The factors that control the migration of lung epithelial cells are incompletely understood. We examined isolated murine type II alveolar epithelial cells (AECs) for expression of CC chemokine receptor 2 (CCR2) and functional consequences of the binding of the main CCR2 ligand monocyte chemoattractant protein-1 (MCP-1). We found that primary AECs bound MCP-1 and expressed CCR2 mRNA. These cells demonstrated functional consequences of CCR2 expression with migration in response to MCP-1 in chemotaxis/haptotaxis assays. Primary AECs cultured from mice lacking CCR2 did not respond to MCP-1. Monolayers of AECs lacking CCR2 demonstrated delayed closure of mechanical wounds compared with AEC monolayers expressing CCR2. Delayed closure of mechanical wounds of wild-type AECs was also demonstrated in the presence of anti-MCP-1 antibody. These data demonstrate for the first time that AECs express CCR2 and are capable of using this receptor for chemotaxis and healing of wounds. CCR2-MCP-1 interactions may be important in the process of reepithelialization after lung injury.     

Anaphylatoxins orchestrate Th17 response via interactions between CD16+ monocytes and pleural mesothelial cells in tuberculous pleural effusion

The complement system is activated in tuberculous pleural effusion (TPE), with increased levels of the anaphylatoxins stimulating pleural mesothelial cells (PMCs) to secrete chemokines, which recruit nonclassical monocytes to the pleural cavity. The differentiation and recruitment of naive CD4⁺ T cells are induced by pleural cytokines and PMC-produced chemokines in TPE. However, it is unclear whether anaphylatoxins orchestrate CD4⁺ T cell response via interactions between PMCs and monocytes in TPE. In this study, CD16⁺ and CD16^- monocytes isolated from TPE patients were cocultured with PMCs pretreated with anaphylatoxins. After removing the PMCs, the conditioned monocytes were cocultured with CD4⁺ T cells. The levels of the cytokines were measured in PMCs and monocyte subsets treated separately with anaphylatoxins. The costimulatory molecules were assessed in conditioned monocyte subsets. Furthermore, CD4⁺ T cell response was evaluated in different coculture systems. The results indicated that anaphylatoxins induced PMCs and CD16⁺ monocytes to secrete abundant cytokines capable of only inducing Th17 expansion, but Th1 was feeble. In addition, costimulatory molecules were more highly expressed in CD16⁺ than in CD16⁻ monocytes isolated from TPE. The interactions between monocytes and PMCs enhanced the ability of PMCs and monocytes to produce cytokines and that of monocytes to express HLA-DR, CD40, CD80 and CD86, which synergistically induced Th17 expansion. In the above process, anaphylatoxins enhanced the interactions between monocytes and PMCs by increasing the level of the cytokines IL-1β, IL-6, IL-23 and upregulating the phenotype of CD40 and CD80 in CD16⁺ monocytes. Collectively, these data indicate that anaphylatoxins play a central role in orchestrating Th17 response mainly via interactions between CD16⁺ monocytes and PMCs in TPE.     

Migration of eosinophils across endothelial cell monolayers: interactions among IL-5, endothelial-activating cytokines, and C-C chemokines

Eosinophils are the predominant cell type recruited in inflammatory reactions in response to allergen challenge. The mechanisms of selective eosinophil recruitment in allergic reactions are not fully elucidated. In this study, the ability of several C-C chemokines to induce transendothelial migration (TEM) of eosinophils in vitro was assessed. Eotaxin, eotaxin-2, monocyte chemotactic protein (MCP)-4, and RANTES induced eosinophil TEM across unstimulated human umbilical vein endothelial cells (HUVEC) in a concentration-dependent manner with the following rank order of potency: eotaxin approximately eotaxin-2 > MCP-4 approximately RANTES. The maximal response induced by eotaxin or eotaxin-2 exceeded that of RANTES or MCP-4. Preincubation of eosinophils with anti-CCR3 Ab (7B11) completely blocked eosinophil TEM induced by eotaxin, MCP-4, and RANTES. Activation of endothelial cells with IL-1beta or TNF-alpha induced concentration-dependent migration of eosinophils, which was enhanced synergistically in the presence of eotaxin and RANTES. Anti-CCR3 also inhibited eotaxin-induced eosinophil TEM across TNF-alpha-stimulated HUVEC. The ability of eosinophil-active cytokines to potentiate eosinophil TEM was assessed by investigating eotaxin or RANTES-induced eosinophil TEM across resting and IL-1beta-stimulated HUVEC in the presence or absence of IL-5. The results showed synergy between IL-5 and the chemokines but not between IL-5 and the endothelial activator IL-1beta. Our data suggest that eotaxin, eotaxin-2, MCP-4, and RANTES induce eosinophil TEM via CCR3 with varied potency and efficacy. Activation of HUVEC by IL-1beta or TNF-alpha or priming of eosinophils by IL-5 both promote CCR3-dependent migration of eosinophils from the vasculature in conjunction with CCR3-active chemokines.     

Lethal (2) giant larvae regulates pleural mesothelial cell polarity in pleural fibrosis

Pleural fibrosis is barely reversible and the underlying mechanisms are poorly understood. Pleural mesothelial cells (PMCs) which have apical-basal polarity play a key role in pleural fibrosis. Loss of cell polarity is involved in the development of fibrotic diseases. Partition defective protein (PAR) complex is a key regulator of cell polarity. However, changes of PMC polarity and PAR complex in pleural fibrosis are still unknown. In this study, we observed that PMC polarity was lost in fibrotic pleura. Next we found increased Lethal (2) giant larvae (Lgl) bound with aPKC and PAR-6B competing against PAR-3A in PAR complex, which led to cell polarity loss. Then we demonstrated that Lgl1 siRNA prevented cell polarity loss in PMCs, and Lgl1 conditional knockout (ER-Cre^+/?Lgl1^flox/flox) attenuated pleural fibrosis in a mouse model. Our data indicated that Lgl1 regulates cell polarity of PMCs, inhibition of Lgl1 and maintenance of cell polarity in PMCs could be a potential therapeutic treatment approach for pleural fibrosis.     

Role of monocyte chemotactic protein-1/CC chemokine ligand 2 on gamma delta T lymphocyte trafficking during inflammation induced by lipopolysaccharide or Mycobacterium bovis bacille Calmette-Guérin

Gammadelta T lymphocytes are involved in a great variety of inflammatory and infectious responses. However, the mechanisms by which gammadelta T lymphocytes migrate to inflamed sites are poorly understood. In this study we investigate the role of monocyte chemotactic protein (MCP)-1 in regulating gammadelta T cell migration after LPS or Mycobacterium bovis bacille Calmette-Guérin (BCG) challenge. LPS-induced gammadelta T cell influx was significantly inhibited by either pretreatment with dexamethasone or vaccinia virus Lister 35-kDa chemokine binding protein, vCKBP, a CC chemokine neutralizing protein, suggesting a role for CC chemokines in this phenomenon. LPS stimulation increased the expression of MCP-1 mRNA and protein at the inflammation site within 6 h. It is noteworthy that LPS was unable to increase MCP-1 production or gammadelta T cell recruitment in C3H/HeJ, indicative of the involvement of Toll-like receptor 4. Gammadelta T cells express MCP-1 receptor CCR2. Pretreatment with anti-MCP-1 mAb drastically inhibited LPS-induced in vivo gammadelta T cell mobilization. Indeed, MCP-1 knockout mice were unable to recruit gammadelta T cells to the pleural cavity after LPS stimulation, effect that could be restored by coadministration of MCP-1. In addition, BCG-induced gammadelta lymphocyte accumulation was significantly reduced in MCP-1 knockout mice when compared with wild-type mice. In conclusion, our results indicate that LPS-induced gammadelta T lymphocyte migration is dependent on Toll-like receptor 4 and sensitive to both dexamethasone and CC chemokine-binding protein inhibition. Moreover, by using MCP-1 neutralizing Abs and genetically deficient mice we show that LPS- and BCG-induced gammadelta T lymphocyte influx to the pleural cavity of mice is mainly orchestrated by the CC chemokine MCP-1.     

TGF-beta1 induces progressive pleural scarring and subpleural fibrosis

Pleural fibrosis is a misunderstood disorder which can cause severe restrictive lung disease with high morbidity and even mortality. The condition can develop in response to a large variety of diseases and tissue injury, among them infectious disease, asbestos, drugs, and radiation therapy. There is no efficient treatment to reverse established pleural fibrosis. TGF-beta1 is suspected, even if not proven, as a key cytokine in this process. In this study, we used adenoviral gene transfer of TGF-beta1 to the pleural mesothelium in rats. We show that local and transient TGF-beta1 overexpression induces homogenous, prolonged, and progressive pleural fibrosis without pleurodesis, associated with severe impairment of pulmonary function. We further demonstrate that pleural fibrosis can expand into the lung parenchyma from the visceral layer, but not into the muscle from the parietal layer. We provide evidence that matrix accumulation and fibrosis within the parenchyma evolved through a process involving "mesothelial-fibroblastoid transformation" and suggest that the pleural mesothelial cell may be an important player involved in the development of the subpleural distribution pattern known to be a hallmark of pulmonary fibrosis. This new model of pleural fibrosis will allow us to better understand the mechanisms of progressive fibrogenesis, and to explore novel antifibrotic therapies in the pleural cavity.     

Innate immune responses in murine pleural mesothelial cells: Toll-like receptor-2 dependent induction of beta-defensin-2 by staphylococcal peptidoglycan

The innate immune response is mediated in part by pattern recognition receptors including Toll-like receptors (TLRs). The pleural mesothelial cells (PMCs) that line the pleural surface are in direct contact with pleural fluid and accordingly carry the risk of exposure to infiltrating microorganisms or their components in an event of a complicated parapneumonic effusion. Here we show that murine primary PMCs constitutively express TLR-1 through TLR-9 and, upon activation with peptidoglycan (PGN), mouse PMC produce antimicrobial peptide beta-defensin-2 (mBD-2). Treatment of PMCs with staphylococcal PGN, a gram-positive bacterial cell wall component and a TLR-2 agonist, resulted in a significant increase in TLR-2 and mBD-2 expression. Silencing of TLR-2 expression by small interfering RNA led to the downregulation of PGN-induced mBD-2 expression, thereby establishing causal relationship between the activation of TLR-2 receptor and mBD-2 production. PMCs exposed to PGN showed increased p38 MAPK activity. In addition, PGN-induced mBD-2 expression was attenuated by SB203580, a p38 MAPK inhibitor, underlining the importance of p38 MAPK in mBD-2 induction. Inhibition of erk1/erk2 or phosphatidylinositol 3-kinase did not block PGN-induced mBD-2 expression in PMC. PGN-activated PMC-derived mBD-2 significantly killed Staphylococcus aureus, and mBD-2-neutralizing antibodies blunted this antimicrobial activity. Taken together, these data indicate that PMCs may contribute to host innate immune defense upon exposure to gram-positive bacteria or their products within the pleural space by upregulating TLR-2 and mBD-2 expression.     

Differentiation and recruitment of Th9 cells stimulated by pleural mesothelial cells in human Mycobacterium tuberculosis infection

Newly discovered IL-9–producing CD4⁺ helper T cells (Th9 cells) have been reported to contribute to tissue inflammation and immune responses, however, differentiation and immune regulation of Th9 cells in tuberculosis remain unknown. In the present study, our data showed that increased Th9 cells with the phenotype of effector memory cells were found to be in tuberculous pleural effusion as compared with blood. TGF-β was essential for Th9 cell differentiation from naïve CD4⁺ T cells stimulated with PMA and ionomycin in vitro for 5 h, and addition of IL-1β, IL-4 or IL-6 further augmented Th9 cell differentiation. Tuberculous pleural effusion and supernatants of cultured pleural mesothelial cells were chemotactic for Th9 cells, and this activity was partly blocked by anti-CCL20 antibody. IL-9 promoted the pleural mesothelial cell repairing and inhibited IFN-γ-induced pleural mesothelial cell apoptosis. Moreover, pleural mesothelial cells promoted Th9 cell differentiation by presenting antigen. Collectively, these data provide new information concerning Th9 cells, in particular the collaborative immune regulation between Th9 cells and pleural mesothelial cells in human M. tuberculosis infection. In particular, pleural mesothelial cells were able to function as antigen-presenting cells to stimulate Th9 cell differentiation.     

Bacterial induction of pleural mesothelial monolayer barrier dysfunction

Pneumonia remains one of the most common infectious causes of mortality. Patients with pneumonia develop parapneumonic effusions with a high neutrophil count as well as high protein concentrations. We hypothesized that pulmonary parenchymal bacterial infection causes a permeability change in the pleural mesothelium by inducing the production of vascular endothelial growth factor (VEGF). Complicated parapneumonic pleural effusions (empyema) have a 19-fold higher VEGF level than pleural fluids secondary to congestive heart failure and a 4-fold higher level than pleural fluids secondary to uncomplicated parapneumonic effusions. We also analyzed the influence of live Staphylococcus aureus on mesothelial barrier function using a model of confluent mesothelial monolayers. There was a significant drop in electrical resistance across S. aureus-infected pleural mesothelial cell (PMC) monolayers. Recombinant VEGF also decreases PMC electrical resistance. Neutralizing antibodies to VEGF significantly inhibited the drop in PMC electrical resistance caused by S. aureus. S. aureus infection also caused a significant increase in protein leak across confluent mesothelial monolayers. Our results suggest that bacterial pathogens induce VEGF release in mesothelial cells and alter mesothelial permeability, leading to protein exudation in empyema.     

MCP-1 deficiency is associated with reduced intimal hyperplasia after arterial injury

Monocyte chemoattractant protein (MCP)-1 is abundant in smooth muscle cells (SMC) and macrophages of atherosclerotic plaques and in the injured arterial wall. MCP-1 and its receptor, CCR2, are important mediators of macrophage accumulation and atherosclerotic plaque progression. We have recently reported that CCR2(-/-) mice have a approximately 60% decrease in intimal hyperplasia and medial DNA synthesis in response to femoral arterial injury. We have now examined the response to femoral arterial injury in MCP-1(-/-) mice. MCP-1 deficiency was associated with a approximately 30% reduction in intimal hyperplasia at 4 weeks and was not associated with diminished medial DNA synthesis. Despite inducing tissue factor in SMC culture, MCP-1 deficiency was not associated with a decrease in neointimal tissue factor after injury. These data suggest that MCP-1 and CCR2 deficiencies have distinct effects on arterial injury. The effects of MCP-1 on intimal hyperplasia may be mediated largely through SMC migration.     

Crosstalk between pleural mesothelial cell and lung fibroblast contributes to pulmonary fibrosis

Idiopathic pulmonary fibrosis (IPF) is a specific form of chronic, progressive and fibrosing interstitial pneumonia of unknown cause. The main feature of IPF is a heterogeneous appearance with areas of sub-pleural fibrosis. However, the mechanism of sub-pleural fibrosis was poorly understood. In this study, our in vivo study revealed that pleural mesothelial cells (PMCs) migrated into lung parenchyma and localized alongside lung fibroblasts in sub-pleural area in mouse pulmonary fibrosis. Our in vitro study displayed that cultured-PMCs-medium induced lung fibroblasts transforming into myofibroblast, cultured-fibroblasts-medium promoted mesothelial-mesenchymal transition of PMCs. Furthermore, these changes in lung fibroblasts and PMCs were prevented by blocking TGF-β1/Smad2/3 signaling with SB431542. TGF-β1 neutralized antibody attenuated bleomycin-induced pulmonary fibrosis. Similar to TGF-β1/Smad2/3 signaling, wnt/β-catenin signaling was also activated in the process of PMCs crosstalk with lung fibroblasts. Moreover, inhibition of CD147 attenuated cultured-PMCs-medium induced collagen-I synthesis in lung fibroblasts. Blocking CD147 signaling also prevented bleomycin-induced pulmonary fibrosis. Our data indicated that crosstalk between PMC and lung fibroblast contributed to sub-pleural pulmonary fibrosis. TGF-β1, Wnt/β-catenin and CD147 signaling was involved in the underling mechanism.     

Basophil responses to chemokines are regulated by both sequential and cooperative receptor signaling

To investigate human basophil responses to chemokines, we have developed a sensitive assay that uses flow cytometry to measure leukocyte shape change as a marker of cell responsiveness. PBMC were isolated from the blood of volunteers. Basophils were identified as a single population of cells that stained positive for IL-3Ralpha (CDw123) and negative for HLA-DR, and their increase in forward scatter (as a result of cell shape change) in response to chemokines was measured. Shape change responses of basophils to chemokines were highly reproducible, with a rank order of potency: monocyte chemoattractant protein (MCP) 4 (peak at <1 nM) >/= eotaxin-2 = eotaxin-3 >/= eotaxin > MCP-1 = MCP-3 > macrophage-inflammatory protein-1alpha > RANTES = MCP-2 = IL-8. The CCR4-selective ligand macrophage-derived chemokine did not elicit a response at concentrations up to 10 nM. Blocking mAbs to CCR2 and CCR3 demonstrated that responses to higher concentrations (>10 nM) of MCP-1 were mediated by CCR3 rather than CCR2, whereas MCP-4 exhibited a biphasic response consistent with sequential activation of CCR3 at lower concentrations and CCR2 at 10 nM MCP-4 and above. In contrast, responses to MCP-3 were blocked only in the presence of both mAbs, but not after pretreatment with either anti-CCR2 or anti-CCR3 mAb alone. These patterns of receptor usage were different from those seen for eosinophils and monocytes. We suggest that cooperation between CCRs might be a mechanism for preferential recruitment of basophils, as occurs in tissue hypersensitivity responses in vivo.     

Induction of low dose oral tolerance in monocyte chemoattractant protein-1- and CCR2-deficient mice

The chemokine monocyte chemoattractant protein-1 (MCP-1) and its receptor CCR2 have been shown to play an important role in the migration and trafficking of macrophages and Th1 effector cells in experimental autoimmune encephalomyelitis. Also, MCP-1 has been reported to regulate oral tolerance induction by inhibition of Th1 cell-related cytokines and by the ability of Abs to MCP-1 to inhibit oral tolerance. This study demonstrates that neither MCP-1 nor its receptor CCR2 is required for the induction of oral tolerance. Mice deletional for either MCP-1 or CCR2 had suppressed cell-proliferative and Th1 responses following oral administration and immunization with myelin oligodendrocyte glycoprotein (MOG(35-55)). TGF-beta was up-regulated in fed and immunized deletional mice, while IL-4 was absent from deletional mice, but up-regulated in controls. Decreased experimental autoimmune encephalomyelitis severity was found in MOG(35-55)-fed MCP-1 deletional mice, indicating induction of oral tolerance. These results demonstrate that MCP-1 is not required for induction of oral tolerance and that MCP-1 and CCR2 are essential for up-regulation of IL-4 in tolerized mice.     

Cell origins and diagnostic accuracy of interleukin 27 in pleural effusions

The objective of the present study was to investigate the presence of interleukin (IL)-27 in pleural effusions and to evaluate the diagnostic significance of pleural IL-27. The concentrations of IL-27 were determined in pleural fluids and sera from 68 patients with tuberculous pleural effusion, 63 malignant pleural effusion, 22 infectious pleural effusion, and 21 transudative pleural effusion. Flow cytometry was used to identify which pleural cell types expressed IL-27. It was found that the concentrations of pleural IL-27 in tuberculous group were significantly higher than those in malignant, infectious, and transudative groups, respectively. Pleural CD4(+) T cells, CD8(+) T cells, NK cells, NKT cells, B cells, monocytes, macrophages, and mesothelial cells might be the cell sources for IL-27. IL-27 levels could be used for diagnostic purpose for tuberculous pleural effusion, with the cut off value of 1,007 ng/L, IL-27 had a sensitivity of 92.7% and specificity of 99.1% for differential diagnosing tuberculous pleural effusion from non-tuberculous pleural effusions. Therefore, compared to non-tuberculous pleural effusions, IL-27 appeared to be increased in tuberculous pleural effusion. IL-27 in pleural fluid is a sensitive and specific biomarker for the differential diagnosing tuberculous pleural effusion from pleural effusions with the other causes.     

ICAM-1 mediated tumor-mesothelial cell adhesion is modulated by IL-6 and TNF-alpha: a potential mechanism by which surgical trauma increases peritoneal metastases

Peritoneal metastases frequently occur in different gastrointestinal cancers and have a poor prognosis. It is known that surgical injury promotes tumor growth and local recurrence rates and also the degree of surgical trauma correlated with the amount of tumor implantation into the peritoneum. The mechanism that mediates tumor cell adhesion to the mesothelium is not fully understood. This study investigates the role of ICAM-1, an important mediator of trans-mesothelial leucocyte migration, in tumor-mesothelial interactions as the initial step in the development of peritoneal recurrence using an in vitro model incorporating mesothelial cell monolayer derived from omental samples. We also investigate how the cytokines interleukins 6 (IL-6) and tumor necrosis factor alpha (TNF-alpha) modulate this process. We demonstrate that ICAM-1 blockade reduces the ability of both pancreatic and colonic cancer cell lines to adhere to the mesothelium. Preincubation of the mesothelial cell monolayer with either IL-6 or TNF-alpha enhances tumor cell adhesion, and this is associated with an increased expression of ICAM-1. Mesothelial CD44 expression, which has previously been implicated in this process, was unaffected by these cytokines. The use of an inhibitory monoclonal antibody against ICAM-1 attenuated the enhanced adhesion mediated by IL-6 or TNF-alpha. This study suggests that mesothelial ICAM-1 plays a role in the adhesion of tumor cells to the peritoneum in the development of peritoneal metastases.     

Mycobacterium tuberculosis Upregulates TNF-α Expression via TLR2/ERK Signaling and Induces MMP-1 and MMP-9 Production in Human Pleural Mesothelial Cells

Background

Tumor necrosis factor (TNF)-α and matrix metalloproteinases (MMPs) are elevated in pleural fluids of tuberculous pleuritis (TBP) where pleural mesothelial cells (PMCs) conduct the first-line defense against Mycobacterium tuberculosis (MTB). However, the clinical implication of TNF-α and MMPs in TBP and the response of PMCs to MTB infection remain unclear.

Methods

We measured pleural fluid levels of TNF-α and MMPs in patients with TBP (n = 18) or heart failure (n = 18) as controls. Radiological scores for initial effusion amount and residual pleural fibrosis at 6-month follow-up were assessed. In vitro human PMC experiments were performed to assess the effect of heat-killed M. tuberculosis H37Ra (MTBRa) on the expression of TNF-α and MMPs.

Results

As compared with controls, the effusion levels of TNF-α, MMP-1 and MMP-9 were significantly higher and correlated positively with initial effusion amount in patients with TBP, while TNF-α and MMP-1, but not MMP-9, were positively associated with residual pleural fibrosis of TBP. Moreover, effusion levels of TNF-α had positive correlation with those of MMP-1 and MMP-9 in TBP. In cultured PMCs, MTBRa enhanced TLR2 and TLR4 expression, activated ERK signaling, and upregulated TNF-α mRNA and protein expression. Furthermore, knockdown of TLR2, but not TLR4, significantly inhibited ERK phosphorylation and TNF-α expression. Additionally, both MTBRa and TNF-α markedly induced MMP-1 and MMP-9 synthesis in human PMCs, and TNF-α neutralization substantially reduced the production of MMP-1, but not MMP-9, in response to MTBRa stimulation.

Conclusion

MTBRa activates TLR2/ERK signalings to induce TNF-α and elicit MMP-1 and MMP-9 in human PMCs, which are associated with effusion volume and pleural fibrosis and may contribute to pathogenesis of TBP. Further investigation of manipulation of TNF-α and MMP expression in pleural mesothelium may provide new insights into the mechanisms and rational treatment strategies for TBP.     

Differential upregulation of chemokine receptors on CD56+ NK cells and their transmigration to the site of infection in tuberculous pleurisy

Chemokines and their receptors orchestrate leukocyte recruitment and confer immunity during Mycobacterium tuberculosis infection. The immunoregulatory and cytotoxic activities of natural killer (NK) cells are essential at the site of infection during tuberculous pleurisy. The frequency, subtypes, and expression of phenotype markers and chemokine receptors on NK cells were assessed by flow cytometry in tuberculous (TB) and nontuberculous (NTB) pleural fluid (PF). Chemotaxis was also shown in response to chemokines. A significant decrease in CD56^dim with no change in CD56^bright NK cells was observed, while a significant increase in activation markers and Toll-like receptors (TLRs) was observed on TB-PF CD56^bright NK cells. Significantly increased expression of chemokine receptors CCR1, CCR2 and CCR7 on CD56^bright and CCR5 on CD56^dim NK cells was observed in the TB group. Transmigration of TB-PF NK cells was significantly high in response to IL-8, IP-10, MCP-1 and SLC. Transmigrated TB-NK cells showed a significant increase in CXCR2, CCR2 and CCR7 expression. The study suggests that CD56^bright NK cells may recognize M. tuberculosis directly using TLRs, HLA-DR and express CD69 as an early activation marker. In addition, CC chemokines induce activation signals in chemokine receptors mediating differential NK cell migration to the site. Thus, NK cells act as first direct sensors and effectors in mycobacterial infection.