In the modern medical era, more diverse and effective treatment options have translated to increased life expectancy. With this increased life span comes increased age-associated disease and the dire need to understand underlying causes so that therapies can be designed to mitigate the burden to health and the economy. Aging exacts a seemingly inevitable multisystem deterioration of function that acts as a risk factor for a variety of age-related disorders, including those that devastate organs of limited regenerative potential, such as the brain. Rather than studying the brain and mechanisms that govern its aging in isolation from other organ systems, an emerging approach is to understand the relatively unappreciated communication that exists between the brain and systemic environment. Revisiting classical methods of experimental physiology in animal models has uncovered surprising regenerative activity in young blood with translational implications for the aging liver, muscle, brain, and other organs. Soluble factors present in young or aged blood are sufficient to improve or impair cognitive function, respectively, suggesting an aging continuum of brain-relevant systemic factors. The age-associated plasma chemokine CCL11 has been shown to impair young brain function while GDF11 has been reported to increase the generation of neurons in aged mice. However, the identities of specific factors mediating memory-enhancing effects of young blood and their mechanisms of action are enigmatic. Here we review brain rejuvenation studies in the broader context of systemic rejuvenation research. We discuss putative mechanisms for blood-borne brain rejuvenation and suggest promising avenues for future research and development of therapies.

CC chemokines, a subfamily of 27 chemotactic cytokines, are a component of intercellular communication, which is crucial for the functioning of the tumor microenvironment. Although many individual chemokines have been well researched, there has been no comprehensive review presenting the role of all known human CC chemokines in the hallmarks of cancer, and this paper aims at filling this gap. The first part of this review discusses the importance of CCL1, CCL3, CCL4, CCL5, CCL18, CCL19, CCL20, CCL21, CCL25, CCL27, and CCL28 in cancer. Here, we discuss the significance of CCL2 (MCP-1), CCL7, CCL8, CCL11, CCL13, CCL14, CCL15, CCL16, CCL17, CCL22, CCL23, CCL24, and CCL26. The presentation of each chemokine includes its physiological function and then the role in tumor, including proliferation, drug resistance, migration, invasion, and organ-specific metastasis of tumor cells, as well as the effects on angiogenesis and lymphangiogenesis. We also discuss the effects of each CC chemokine on the recruitment of cancer-associated cells to the tumor niche (eosinophils, myeloid-derived suppressor cells (MDSC), tumor-associated macrophages (TAM), tumor-associated neutrophils (TAN), regulatory T cells (T_reg)). On the other hand, we also present the anti-cancer properties of CC chemokines, consisting in the recruitment of tumor-infiltrating lymphocytes (TIL).

Keywords: CC chemokine; MCP-1; angiogenesis; anti-cancer therapy; cancer; chemokine; lymphangiogenesis; organ-specific metastasis; tumor; tumor microenvironment.

Cell migration is mediated by a group of chemotactic cytokines called chemokines: low molecular weight molecules that have been shown as important leukocyte chemical attractants to sites of inflammation and infection. Eotaxin-1, also called CCL11, was first described in 1994, as a highly specific eosinophils chemokine. Many cell types including lymphocytes, macrophages, bronchial smooth muscle cells, endothelial cells and eosinophils, are able to produce this chemokine, predominantly after cytokine stimulation, however little is known about its expression in human skin in vivo. Eotaxin-1 also regulates the chemiotaxis and, in some conditions, activation of basophils, mast cells and T lymphocytes. Chemokine receptors are named from their ligand families, thus the CC chemokine eotaxin-1 binds to the CCR3 receptor which is expressed on eosinophis, mast cells, Th2 type lymphocytes and even on keratinocytes. It seems that eotaxin-1 is one of the most important cytokines involved in tissue inflammation playing a central role in the pathogenesis of allergic airway diseases (asthma and rhinitis), in inflammatory bowel disease and gastrointestinal allergic hypersensitivity and recently it has been proposed as a therapeutical target for these conditions. Our group has studied the role of eotaxin-1 in the pathogenesis of two skin conditions: dermatitis herpetiformis and AIDS-associated eosinophilic folliculitis, demonstrating that this chemokine, together with Th2 type cytokines (IL-13 and IL-4) is important in cell recruitment, inflammation and tissue damage; moreover eotaxin has proven to paly an important role in other skin conditions such as, bullous pemphigoid, pemphigoid gestationis, atopic dermatitis and allergic drug reactions Recent advances in the understanding of eotaxin-1-mediated mechanisms of chemotaxis in allergic and inflammatory conditions may predict that therapeutic antagonism is achievable. This paper will focus on the role that eotaxin and its receptor play in the pathogenetical mechanism in a number of dermatologic diseases, some of which, like atopic dermatitis, may benefit from the introduction of novel and more selective therapeutic options.

Cytokines play important roles as regulators of cell functions, and over the last decades a number of cytokine assays have been developed. The aim of the present study was to investigate the effects of age and gender on a large number of cytokines. Plasma samples were collected from 33 healthy blood donors. The samples were analyzed using a multiplex proximity extension assay (PEA) allowing simultaneous measurement of 92 cytokines and four technical controls. Biomarkers with less than 80% quantitative results were excluded leaving 63 cytokines that were analyzed for the effects of gender and age. The plasma level of three of the investigated biomarkers (DNER, MCP-4 and MMP-10) were found to be significantly different for the two genders (adjusted p-value<0.05), and 15 of the biomarkers (CCL11, CCL25, CDCP1, CSF-1, CXCL11, CXCL9, FGF-23, Flt3L, HGF, IL-10RB, MCP-3, MCP-4, MMP-10, OPG, VEGF-A) were significantly associated with age. This study reveals the effects of age and gender on a large number of cytokine assays. CXCL5 and TNFB were significantly higher in females, while the other markers with significant gender-dependent differences were higher in males. For the markers that were significantly associated with age, only CXCL6 was found to decrease with age, while the other biomarkers increased with age.

For decades in China, the Yin-Huang-Qing-Fei capsule (YHQFC) has been widely used in the treatment of chronic bronchitis, with good curative effects. Owing to the complexity of traditional Chinese herbal formulas, the pharmacological mechanism of YHQFC remains unclear. To address this problem, a network pharmacology-based strategy was proposed in this study. At first, the putative target profile of YHQFC was predicted using MedChem Studio, based on structural and functional similarities of all available YHQFC components to the known drugs obtained from the DrugBank database. Then, an interaction network was constructed using links between putative YHQFC targets and known therapeutic targets of chronic bronchitis. Following the calculation of four topological features (degree, betweenness, closeness, and coreness) of each node in the network, 475 major putative targets of YHQFC and their topological importance were identified. In addition, a pathway enrichment analysis based on the Kyoto Encyclopedia of Genes and Genomes pathway database indicated that the major putative targets of YHQFC are significantly associated with various pathways involved in anti-inflammation processes, immune responses, and pathological changes caused by asthma. More interestingly, eight major putative targets of YHQFC (interleukin [IL]-3, IL-4, IL-5, IL-10, IL-13, FCER1G, CCL11, and EPX) were demonstrated to be associated with the inflammatory process that occurs during the progression of asthma. Finally, a molecular docking simulation was performed and the results exhibited that 17 pairs of chemical components and candidate YHQFC targets involved in asthma pathway had strong binding efficiencies. In conclusion, this network pharmacology-based investigation revealed that YHQFC may attenuate the inflammatory reaction of chronic bronchitis by regulating its candidate targets, which may be implicated in the major pathological processes of the asthma pathway.

Ticks secrete a cocktail of immunomodulatory molecules in their saliva during blood-feeding, including chemokine-binding factors that help control the activity of host immunocompetent cells. Here we demonstrate differential dynamics of anti IL-8 (CXCL8), MCP-1 (CCL2), MIP-1 (CCL3), RANTES (CCL5) and eotaxin (CCL11) activities in salivary gland extracts of adult Amblyomma variegatum. Unfed male and female ticks showed activity against all the chemokines except CCL5; anti-CCL11 activity was particularly high. However, during feeding the dynamics of anti-chemokine activity differed significantly between males and females, and varied between chemokines. In males, anti-chemokine activities increased, whereas in females they declined or increased slightly as feeding progressed. The exception was anti-CCL11 activity, which declined and then increased in both males and females. Comparison of salivary gland equivalents of individual ticks prepared at various feeding intervals revealed some differences that were most pronounced between individual females fed for 8 days. These observations reflect the feeding behaviour of male and female A. variegatum. They support the concept of 'mate guarding', in which males help their mates to engorge by controlling their host's immune response, and the possibility that ticks benefit from feeding together by exploiting molecular individuality.

The function of chemokine receptors on structural cells is only partially known. We previously reported the expression of a functional CCR3 receptor on airway epithelial cells (EC). We speculated that CCR3 might drive wound repair and expression of inflammatory genes in epithelium. The human airway EC lines BEAS-2B, 16-HBE, and primary bronchial EC were used to test the effect of in vitro challenge with the CCR3 ligands CCL11/eotaxin, CCL24/eotaxin-2, or CCL26/eotaxin-3 on 1) wound repair, using an established wound model; 2) cell proliferation and chemotaxis, using specific fluorometric assays; and 3) gene expression, using pathway-specific arrays for inflammatory and profibrotic cytokines, chemokines, and chemokine receptor genes. Agonist specificity was tested by cell pretreatment with an AstraZeneca CCR3 antagonist (10(-8) - 10(-6) M). CCL24 challenge significantly accelerated epithelial wound closure, with similar effects exerted by CCL11 and CCL26. This effect was time dependent, submaximal at 1 nM, and comparable in potency to epidermal growth factor. CCL24 induced a concentration-dependent increase in EC proliferation and chemotaxis, with significant effects observed at 10 nM. The AstraZeneca compound selectively inhibited these CCL24-mediated responses. CCL11 induced the up-regulation of several profibrogenic molecules such as fibroblast growth factor 1 and 5 and of several CC and CXC chemokines. Epithelial immunostaining for CCR3 was stronger in bronchial biopsies of asthmatics displaying marked inflammatory changes than in nondiseased samples. Epithelial CCR3 participates in key functions for wound repair, amplifies the expression of profibrogenic and chemokine transcripts, and appears up-regulated in inflamed asthmatic airways.

Mice homozygous for the STAT4-null mutation were sensitized to cockroach Ag, challenged intratracheally 21 days later, and compared with STAT4-competent allergic mice. The STAT4(-/-) mice showed significant decreases in airway hyperreactivity (AHR) and peribronchial eosinophils compared with wild-type controls. In addition, pulmonary levels of chemokines were decreased in the STAT4(-/-) mice, including CC chemokine ligand (CCL)5, CCL6, CCL11, and CCL17. However, levels of Th2-type cytokines, such as IL-4 and IL-13, as well as serum IgE levels were similar in the two groups. Transfer of splenic lymphocytes from sensitized wild-type mice into sensitized STAT4(-/-) mice did not restore AHR in the mutant mice. Furthermore, chemokine production and peribronchial eosinophilia were not restored during the cellular transfer experiments. Thus, it appears that STAT4 expression contributes to a type 2 process such as allergen-induced chemokine production and AHR. In additional studies, competent allergic mice were treated with anti-IL-12 locally in the airways at the time of allergen rechallenge. These latter studies also demonstrated a decrease in AHR. Altogether, these data suggest that STAT4-mediated pathways play a role locally within the airway for the exacerbation of the allergen-induced responses.

Eotaxin-3 (CCL26) is a CC chemokine that signals exclusively via the CCR3 receptor and has eosinophil-selective chemoattractant activity. Comparison of Eotaxin-1 (CCL11) and Eotaxin-2 (CCL24), demonstrates differences in their expression profiles, cell specificity and effector kinetics, implying distinct biological actions. But little data in this regard have been reported for Eotaxin-3. We aimed to analyse the effect of Th2 cytokines and glucocorticoids on Eotaxin-3 mRNA expression in human lung epithelial cells and dermal fibroblasts; cells implicated in the pathogenesis of allergic asthma and allergic dermatitis respectively. Eotaxin-3 mRNA levels in primary dermal fibroblasts and NCI-H727 lung epithelial cells were determined by Northern hybridization. In contrast to Eotaxin-1, Eotaxin-3 mRNA expression was not detected in unstimulated cells. The Th2 cytokines IL-4 and IL-13 induced Eotaxin-3 expression in a time and dose dependent manner, with IL-4 demonstrating a 100-fold greater potency. Unlike Eotaxin-1, Eotaxin-3 mRNA expression was not induced by either tumour necrosis factor (TNF)-alpha or interleukin (IL)-1 beta alone. Both IL-4 and IL-13 acted synergistically with TNF-alpha in superinducing Eotaxin-3 mRNA expression. Dexamethasone pre-treatment diminished induction of Eotaxin-3 mRNA expression. We conclude that modulation of Eotaxin-3 mRNA expression by Th(2) cytokines is different from that of Eotaxin-1 and Eotaxin-2, further supporting a distinct biological role for Eotaxin-3.

Despite curative locoregional treatments for hepatocellular carcinoma (HCC), tumour recurrence rates remain high. The current study was designed to assess the safety and bioactivity of infusion of dendritic cells (DCs) stimulated with OK432, a streptococcus-derived anti-cancer immunotherapeutic agent, into tumour tissues following transcatheter hepatic arterial embolization (TAE) treatment in patients with HCC. DCs were derived from peripheral blood monocytes of patients with hepatitis C virus-related cirrhosis and HCC in the presence of interleukin (IL)-4 and granulocyte-macrophage colony-stimulating factor and stimulated with 0·1 KE/ml OK432 for 2 days. Thirteen patients were administered with 5 × 10⁶ of DCs through arterial catheter during the procedures of TAE treatment on day 7. The immunomodulatory effects and clinical responses were evaluated in comparison with a group of 22 historical controls treated with TAE but without DC transfer. OK432 stimulation of immature DCs promoted their maturation towards cells with activated phenotypes, high expression of a homing receptor, fairly well-preserved phagocytic capacity, greatly enhanced cytokine production and effective tumoricidal activity. Administration of OK432-stimulated DCs to patients was found to be feasible and safe. Kaplan-Meier analysis revealed prolonged recurrence-free survival of patients treated in this manner compared with the historical controls (P = 0·046, log-rank test). The bioactivity of the transferred DCs was reflected in higher serum concentrations of the cytokines IL-9, IL-15 and tumour necrosis factor-α and the chemokines CCL4 and CCL11. Collectively, this study suggests that a DC-based, active immunotherapeutic strategy in combination with locoregional treatments exerts beneficial anti-tumour effects against liver cancer.

In aged mice, we assessed how intensive exercise affects brain bioenergetics, inflammation, and neurogenesis-relevant parameters. After 8 weeks of a supra-lactate threshold treadmill exercise intervention, 21-month-old C57BL/6 mice showed increased brain peroxisome proliferator-activated receptor gamma coactivator-1α protein, mammalian target of rapamycin and phospho-mammalian target of rapamycin protein, citrate synthase messenger RNA, and mitochondrial DNA copy number. Hippocampal vascular endothelial growth factor A (VEGF-A) gene expression trended higher, and a positive correlation between VEGF-A and PRC messenger RNA levels was observed. Brain doublecortin, brain-derived neurotrophic factor, tumor necrosis factor-α, and CCL11 gene expression, as well as plasma CCL11 protein levels, were unchanged. Despite these apparent negative findings, a negative correlation between plasma CCL11 protein levels and hippocampal doublecortin gene expression was observed; further analysis indicated exercise may mitigate this relationship. Overall, our data suggest supra-lactate threshold exercise activates a partial mitochondrial biogenesis in aged mice, and a gene (VEGF-A) known to support neurogenesis. Our data are consistent with another study that found systemic inflammation in general, and CCL11 protein specifically, suppresses hippocampal neurogenesis. Our study supports the view that intense exercise above the lactate threshold may benefit the aging brain; future studies to address the extent to which exercise-generated lactate mediates the observed effects are warranted.

GM-CSF plays an important role in inflammation by promoting the production, activation, and survival of granulocytes and macrophages. In this study, GM-CSF knockout (GM-CSF(-/-)) mice were used to investigate the role of GM-CSF in a model of allergic airway inflammation. In allergic GM-CSF(-/-) mice, eosinophil recruitment to the airways showed a striking pattern, with eosinophils present in perivascular areas, but almost completely absent in peribronchial areas, whereas in wild-type mice, eosinophil infiltration appeared in both areas. In the GM-CSF(-/-) mice, mucus production in the airways was also reduced, and eosinophil numbers were markedly reduced in the bronchoalveolar lavage (BAL)(3) fluid. IL-5 production was reduced in the lung tissue and BAL fluid of GM-CSF(-/-) mice, but IL-4 and IL-13 production, airway hyperresponsiveness, and serum IgE levels were not affected. The presence of eosinophils in perivascular but not peribronchial regions was suggestive of a cell migration defect in the airways of GM-CSF(-/-) mice. The CCR3 agonists CCL5 (RANTES) and CCL11 (eotaxin-1) were expressed at similar levels in GM-CSF(-/-) and wild-type mice. However, IFN-gamma mRNA and protein were increased in the lung tissue and BAL fluid in GM-CSF(-/-) mice, as were mRNA levels of the IFN-gamma-inducible chemokines CXCL9 (Mig), CXCL10 (IP-10), and CXCL11 (I-Tac). Interestingly, these IFN-gamma-inducible chemokines are natural antagonists of CCR3, suggesting that their overproduction in GM-CSF(-/-) mice contributes to the lack of airway eosinophils. These findings demonstrate distinctive abnormalities to a model of allergic asthma in the absence of GM-CSF.

Cantharidin skin blisters were examined over two days to model the acute and resolving phases of inflammation in human skin. Four blisters were created by topical administration of cantharidin (0.1% v/v) to the forearm of healthy volunteers, with IRB approval. Duplicate skin blisters were aspirated at 16 and 40 hours to model the proinflammatory and resolving phases, respectively. There was a significant increase in leukocyte infiltrate at 40 h with appearance of a "resolving macrophage" phenotype CD14(+)CD163(+) by flow cytometry. Neutrophils acquired apoptotic markers at 40 h and were observed to be phagocytosed by macrophagic "Reiter's" cells. Multiplex cytokine analysis demonstrated that monocyte chemoattractant protein (MCP-1/CCL2), interleukin- (IL-) 6, IL-8/CXCL8, macrophage inflammatory protein (MIP1 α /CCL3), MIP-1 β /CCL4, tumor necrosis factor- (TNF-) α , and eotaxin (CCL11) were all significantly upregulated at 16 h compared with 40 h. In contrast, immunoregulatory transforming growth factor- (TGF-) β , macrophage-derived chemokine (MDC/CCL22), and interferon-inducible protein (IP-10/CXCL10) were significantly elevated at 40 h. Our results demonstrate that the phases of inflammation and resolution can be discriminated in a two-day model of dermal wound healing. This confirms and extends our understanding of wound repair in humans and provides a powerful research tool for use in clinical settings and to track the molecular benefits of therapeutic intervention.

Post-translational modification of chemokines is an essential regulatory mechanism to enhance or dampen the inflammatory response. CD26/dipeptidylpeptidase IV, ubiquitously expressed in tissues and blood, removes NH2-terminal dipeptides from proteins with a penultimate Pro or Ala. A large number of human chemokines, including CXCL2, CXCL6, CXCL9, CXCL10, CXCL11, CXCL12, CCL3L1, CCL4, CCL5, CCL11, CCL14, and CCL22, are cleaved by CD26; however, the efficiency is clearly influenced by the amino acids surrounding the cleavage site and although not yet proven, potentially affected by the chemokine concentration and interactions with third molecules. NH2-terminal cleavage of chemokines by CD26 has prominent effects on their receptor binding, signaling, and hence, in vitro and in vivo biologic activities. However, rather than having a similar result, the outcome of NH2-terminal truncation is highly diverse. Either no difference in activity or drastic alterations in receptor recognition/specificity and hence, chemotactic activity are observed. Analogously, chemokine-dependent inhibition of HIV infection is enhanced (for CCL3L1 and CCL5) or decreased (for CXCL12) by CD26 cleavage. The occurrence of CD26-processed chemokine isoforms in plasma underscores the importance of the in vitro-observed CD26 cleavages. Through modulation of chemokine activity, CD26 regulates leukocyte/tumor cell migration and progenitor cell release from the bone marrow, as shown by use of mice treated with CD26 inhibitors or CD26 knockout mice. As chemokine processing by CD26 has a significant impact on physiologic and pathologic processes, application of CD26 inhibitors to affect chemokine function is currently explored, e.g., as add-on therapy in viral infection and cancer.

Dipeptidyl peptidase IV (DP-IV/CD26), fibroblast activation protein (FAP), DP-like 1 (DPL1), DP8, DP9, and DPL2 comprise the CD26 gene family. CD26/DP-IV has roles in liver disease, T cell costimulation, chemokine biology, type II diabetes, and tumor biology. DPIV substrates include the glucagonlike peptides, neuropeptide Y, and the chemokines CCL3, CCL5, CCL11, CCL22, and CXCL12. We have proposed that the extracellular region of CD26 is analogous to prolyl oligopeptidase in consisting of an alpha/beta hydrolase domain contributed by both N- and C-terminal portions of the polypeptide and a seven-blade beta-propeller domain. Replacing the C-terminal portion of the predicted alpha/beta hydrolase domain of CD26 (residues 501-766) with the homologous portion of DP8 or DP9 produced intact proteins. However, these chimeric proteins lacked dimerization and peptidase activity, suggesting that CD26 dimerization requires the C-terminal portion of the alpha/beta hydrolase domain. Deleting some N-terminal residues of the alpha/beta hydrolase domain of CD26 ablated peptidase activity and greatly diminished cell surface expression. Together with previous data that CD26 peptidase activity requires the C-terminal 20 residues, this suggests that peptidase activity requires the entire alpha/beta hydrolase domain. The catalytic triad of DP8 was shown to be Ser(739)-Asp (817)-His(849). Glu(259) of DP8, a residue distant from the catalytic triad yet greatly conserved in the CD26 gene family, was shown to be required for peptidase activity. These data concord with our predicted CD26 structure, indicate that biosynthesis of a functional fragment of CD26 is difficult, and confirm the functional homology of DP8 with CD26.

Previous studies have shown that the myeloid-specific deficiency of the transcription factor Krüppel-like factor 2 (KLF2) accelerates atherosclerosis in hypercholesterolemic Ldlr(-/-) mice due to the enhanced adhesion of myeloid cells to activated endothelial cells in the vessel wall. This study revealed elevated basal inflammation with elevated plasma levels of Ccl2, Ccl4, Ccl5, and Ccl11 in the myeloid-specific KLF2 knock-out (myeKlf2(-/-)) mice. Peritoneal macrophages isolated from myeKlf2(-/-) mice showed increased mRNA levels of several inflammatory mediators, including Ccl2, Ccl5, Ccl7, Cox-2, Cxcl1, and IL-6. In contrast, the levels of two microRNAs, miR-124a and miR-150, were lower in Klf2(-/-) macrophages compared with Klf2(+/+) macrophages. Additional studies showed a direct inverse relationship between miR-124a levels with Ccl2 expression, with anti-miR-124a increasing Ccl2 mRNA levels in Klf2(+/+) macrophages, whereas the restoration of miR-124a levels in Klf2(-/-) macrophages significantly reduced Ccl2 mRNA expression. Likewise, the inverse relationship was observed between miR-150 levels and Cxcl1 expression in Klf2(+/+) and Klf2(-/-) mice. Moreover, miR150 likely regulates the miR124a expression and thus augments expression of inflammatory mediators in myeKlf2(-/-) macrophages. This study documented that the transcription factor KLF2 modulates inflammatory chemokine production via regulation of microRNA expression levels in immune cells.

CCL11/eotaxin-1 is a potent eosinophilic CC chemokine expressed by primary human fibroblasts. The combination of TGF-β1 and IL-13 synergistically increases CCL11 expression, but the mechanisms behind the synergy are unclear. To address this, human airway fibroblast cultures from normal and asthmatic subjects were exposed to IL-13 alone or TGF-β1 plus IL-13. Transcriptional (nuclear run-on) and posttranscriptional (mRNA stability) assays confirmed that transcriptional regulation is critical for synergistic expression of CCL11. TGF-β1 plus IL-13 synergistically increased STAT-6 phosphorylation, nuclear translocation, and binding to the CCL11 promoter as compared with IL-13 alone. STAT-6 small interfering RNA significantly knocked down both STAT-6 mRNA expression and phosphorylation and inhibited CCL11 mRNA and protein expression. Regulation of the IL-4Rα complex by TGF-β1 augmented IL-13 signaling by dampening IL-13Rα2 expression, overcoming IL-13's autoregulation of its pathway and enhancing the expression of CCL11. Our data suggest that TGF-β1 induced activation of the MEK/ERK pathway reduces IL-13Rα2 expression induced by IL-13. Thus, TGF-β1, a pleiotropic cytokine upregulated in asthmatic airways, can augment eosinophilic inflammation by interfering with IL-13's negative feedback autoregulatory loop under MEK/ERK-dependent conditions.

The regenerative capacity of peripheral nerves declines during aging, contributing to the development of neuropathies, limiting organism function. Changes in Schwann cells prompt failures in instructing maintenance and regeneration of aging nerves; molecular mechanisms of which have yet to be delineated. Here, we identified an altered inflammatory environment leading to a defective Schwann cell response, as an underlying mechanism of impaired nerve regeneration during aging. Chronic inflammation was detected in intact uninjured old nerves, characterized by increased macrophage infiltration and raised levels of monocyte chemoattractant protein 1 (MCP1) and CC chemokine ligand 11 (CCL11). Schwann cells in the old nerves appeared partially dedifferentiated, accompanied by an activated repair program independent of injury. Upon sciatic nerve injury, an initial delayed immune response was followed by a persistent hyperinflammatory state accompanied by a diminished repair process. As a contributing factor to nerve aging, we showed that CCL11 interfered with Schwann cell differentiation in vitro and in vivo. Our results indicate that increased infiltration of macrophages and inflammatory signals diminish regenerative capacity of aging nerves by altering Schwann cell behavior. The study identifies CCL11 as a promising target for anti-inflammatory therapies aiming to improve nerve regeneration in old age.

BACKGROUND

Chemokines are a class of cytokines with chemotactic properties shown to be induced by M. tuberculosis or its antigens in vitro and in experimental infection in vivo. A few studies have also demonstrated the expression of chemokines in clinical samples of patients with active tuberculosis (TB). In the present work, we measured the concentration of chemokines in plasma samples of HIV-negative patients with pulmonary tuberculosis at different stages of chemotherapy. For comparison, we also evaluated the levels of sTNFR1 and TNF-alpha.

METHODS

Cytokines and chemokines were measured by ELISA in healthy individuals and patients with active pulmonary TB at different stages of treatment.

RESULTS

The concentrations of CXCL8, CXCL9 and sTNFR1 were elevated in patients with active pulmonary TB but returned to background levels at 4-6 months of chemotherapy. The concentration of CCL11 was elevated in patients with active pulmonary tuberculosis when compared to control and remained elevated throughout the specific therapy. There was no difference in the plasma concentration of CCL2 and CXCL10 between pulmonary TB patients and control subjects.

CONCLUSIONS

Measurement of the CXCL8, CXCL9 and sTNFR1 may be useful to assess response to treatment in pulmonary TB patients.

Asthma is a common disease that causes considerable morbidity. Increased numbers of airway eosinophils are a hallmark of asthma. Mechanisms controlling the entry of eosinophils into asthmatic lung have been intensively investigated, but factors regulating migration within the tissue microenvironment are less well understood. We modeled this by studying chemoattractant and growth factor-mediated human eosinophil migration within a three-dimensional collagen matrix. Stimulation with GM-CSF induced dose-dependent, random migration with a maximum of 77 +/- 4.7% of cells migrating. In contrast, CCL11 and C5a caused a more modest although significant degree of migration (19 +/- 1.8% and 20 +/- 2.6%, respectively). Migration to GM-CSF was partially dependent on Ca(2+) and alpha(M)beta(2) integrins. The Rho family of small GTPases regulates intracellular signaling of cell migration. GM-CSF-induced migration was only partially dependent on Rho kinase/Rho-associated kinase (ROCK) and was independent of RhoA activation. In contrast, CCL11-induced migration was fully dependent on both RhoA and ROCK. Activation of RhoA was therefore neither necessary nor sufficient to cause eosinophil migration in a three-dimensional collagen environment. This study suggests that eosinophil growth factors are likely to be required for eosinophil migration within the bronchial mucosa, and this involves signal transduction pathways distinct from those used by G protein-associated chemoattractants.

BACKGROUND

Circulating cytokine patterns may be relevant for the diagnosis of asthma, for the discrimination of certain phenotypes, and prognostic factors for exacerbation of disease.

RESULTS

In this study we investigated serum samples from 944 individuals of 218 asthma-affected families by a multiplex, microsphere based system detecting at high sensitivity eleven asthma associated mediators: eotaxin (CCL11), granulocyte macrophage stimulating factor (GM-CSF), interferon gamma (IFNγ), interleukin-4 (IL-4), IL-5, IL-8, IL-10, IL-12 (p40), IL-13, IL-17 and tumor necrosis factor alpha (TNFα). Single cytokine levels were largely similar between asthmatic and healthy individuals when analysing asthma as single disease entity. Regulatory differences between parental and pediatric asthma were reflected by six of the eleven mediators analyzed (eotaxin, IL-4, IL-5, IL-10, IL-12, TNFα). IL-12 (p40) and IL-5 were the best predictor for extrinsic asthma in children with an increased odds ratio of 2.85 and 1.96 per log pg/ml increase (IL-12 (p40): 1.2-6.8, p=0.019, and IL-5: 1.2-2.5, p=0.025). Frequent asthma attacks in children are associated with elevated IL-5 serum levels (p=0.013). Cytokine patterns seem to be individually balanced in both, healthy and diseased adults and children, with various cytokines correlating among each other (IL-17 and IFNγ (rs=0.67), IL-4 and IL-5 (rs=0.55), IFNγ and GM-CSF (rs=0.54)).

CONCLUSIONS

Our data support mainly an age- but also an asthma phenotype-dependent systemic immune regulation.

In the present study, we investigated the regulation of chemokine-mediated responses and receptor expression on eosinophils from mice. MIP-1alpha (CCL3) and eotaxin (CCL11) induced a significant and only partially overlapping intracellular calcium flux in antigen-elicited and peripheral blood eosinophils, and MCP-1 (CCL2), MDC (CCL22), MIP-1beta (CCL4), and TCA-3 (CCL1) did not. To demonstrate functional use of the specific receptors, we examined chemotactic responses. Peripheral blood eosinophils migrated toward MIP-1alpha (CCL3) and eotaxin (CCL11) but not MCP-1 (CCL2), MDC (CCL22), MIP-1beta (CCL4), and TCA-3 (CCL1). Antigen-elicited eosinophils migrated toward MIP-1alpha (CCL3) and eotaxin (CCL11), but also migrated in response to MIP-1beta (CCL4) and TCA-3 (CCL1), suggesting the up-regulation of additional chemokine receptors on antigen-elicited eosinophils. The up-regulation of the additional chemokine-receptor responses appeared to be in part because of cytokine activation, because TNF-alpha and/or IL-4 were able to up-regulate CCR1, -3, -5, and -8 mRNA expression in eosinophils as well as migration responses to the appropriate ligands. Using antibodies specific for CCR5 and CCR8, the chemotactic response to MIP-1beta and TCA-3, respectively, was reduced significantly. Finally, the expression of these new receptors appears to have an effect on activation and degranulation because MIP-1beta (CCL4) and TCA-3 (CCL1) induce significant levels of LTC4 from elicited eosinophils. These results suggest that eosinophils may up-regulate and use additional chemokine receptors during progression of inflammatory, allergic responses for migration and activation.

During allergic reactions, basophils migrate from the blood compartment to inflammatory sites, where they act as effector cells in concert with eosinophils. Because transendothelial migration (TEM) represents an essential step for extravasation of cells, for the first time we have studied basophil TEM using HUVEC. Treatment of HUVEC with IL-1beta significantly enhanced basophil TEM, which was further potentiated by the presence of a CCR3-specific ligand, eotaxin/CCL11. In addition to CCR3 ligands, MCP-1/CCL2 was also active on basophil TEM. Although stromal cell-derived factor-1/CXCL12, a CXCR4 ligand, failed to induce TEM in freshly isolated basophils, it caused strong TEM in 24-h cultured cells. IL-3 enhanced basophil TEM by increasing the chemokinetic response. Spontaneous TEM across activated HUVEC was inhibited by treatment of cells with anti-CD18 mAb, but not with anti-CD29 mAb, and also by treatment of HUVEC with anti-ICAM-1 mAb. Anti-VCAM-1 mAb alone failed to inhibit TEM, but showed an additive inhibitory effect in combination with anti-ICAM-1 mAb. In contrast, eotaxin- and IL-3-mediated TEM was significantly inhibited by anti-CD29 mAb as well as anti-CD18 mAb. These results indicate that beta2 integrins play the primary role in basophil TEM, but beta1 integrins are also involved, especially in TEM of cytokine/chemokine-stimulated basophils. In conclusion, the regulatory profile of basophil TEM is very similar to that reported for eosinophils. Our results thus support the previous argument for a close relationship between basophils and eosinophils and suggest that the in vivo kinetics of these two cell types are similar.