Camel whey protein improves lymphocyte function and protects against diabetes in the offspring of diabetic mouse dams.

The prevalence of health problems in the offspring of pregnant diabetic mothers has recently been verified. Therefore, the present study was designed to investigate the influence of dietary camel whey protein (CWP), administered as a supplement to streptozotocin (STZ)-induced diabetic pregnant mice, on the efficiency of the immune system of the offspring. Three groups of female mice (n = 10) were used: non-diabetic control mice, diabetic mice, and diabetic mice orally administered CWP during the pregnancy and lactation periods. We then tested the immune response of B and T cells in adult male offspring (n = 15 in each group) by using flow cytometry, western blotting, and ELISAs. Our data demonstrated that the offspring of diabetic dams exhibited several postpartum complications, such as significant aberrant overexpression of activating transcription factor-3 (ATF-3), significant elevation of the plasma levels of pro-inflammatory cytokines (IL-1ß, IL-6, and TNF-α) and reactive oxygen species (ROS), marked decreases in the plasma levels of IL-2 and IL-7, significant inhibition of CCL21- and CXCL12-mediated chemotaxis of B- and T-lymphocytes, and a marked decrease in the proliferative capacity of antigen-stimulated B- and T-lymphocytes. Interestingly, administration of CWP to diabetic dams substantially restored the expression of ATF-3 and the levels of ROS, pro-inflammatory cytokines, IL-2, and IL-7 in the offspring. Furthermore, the chemotaxis of B- and T-lymphocytes toward CCL21 and CXCL12 and the proliferative capacities of these lymphocytes were restored in the male offspring of diabetic mice administered CWP. Our data provide evidence of a protective role of CWP in decreasing the tendency of the offspring of diabetic mothers to develop diabetes and related complications.

Baicalin attenuates inflammation in mice with OVA-induced asthma by inhibiting NF-κB and suppressing CCR7/CCL19/CCL21.

Baicalin, extracted and purified from the Chinese medicinal plant, Scutellaria baicalensis Georgi (Huang qin in Chinese), exhibits potent anti-inflammatory activity against asthma. However, it remains unknown whether baicalin inhibits the activity of CC chemokine receptor 7 (CCR7) and its ligands, which are crucial for the initiation of airway inflammation. In the present study, we investigated the effects of baicalin on CCR7 and its ligands, CCL19 and CCL21, as well as on the nuclear factor-κB (NF-κB) pathway in a mouse model of asthma. A mouse model of acute asthma was established by exposing the mice to ovalbumin (OVA) (by intraperitoneal injection and inhalational challenge). Within 24 h of the final OVA challenge, lung function was detected by direct airway resistance analysis. Lung tissues were examined for pathological changes. Inflammatory cell counts in bronchoalveolar lavage fluid (BALF) were assessed. ELISA was utilized to evaluate the OVA-IgE, CCL19 and CCL21 levels in BALF. The interleukin (IL)-6 and tumor necrosis factor (TNF)-α levels in serum were also detected by ELISA. The protein expression levels of CCR7, as well as that of phosphorylated IκBα (p-IκBα) and phosphorylated p65 (p-p65) were determined by western blot analysis and RT-qPCR was used to determine the CCR7 mRNA levels. Our data demonstrated that the oral administration of baicalin significantly improved pulmonary function and attenuated inflammatory cell infiltration into the lungs. Baicalin also decreased the levels of OVA-IgE, IL-6, TNF-α and CCR7, as well as those of its ligand, CCL19; the levels of NF-κB were also markedly suppressed by baicalin. The CCR7 mRNA level was substantially decreased. Our results thus suggest that baicalin exerts an inhibitory effect on airway inflammation, and this effect may be associated with the inhibition of CCR7 and CCL19/CCL21, which may provide new mechanistic insight into the anti­inflammatory effects of baicalin.

Oral supplementation of diabetic mice with propolis restores the proliferation capacity and chemotaxis of B and T lymphocytes towards CCL21 and CXCL12 by modulating the lipid profile, the pro-inflammatory cytokine levels and oxidative stress.

BACKGROUND: Type 1 diabetes mellitus (T1D) is a chronic autoimmune disease caused by the selective destruction of pancreatic ß cells, followed by hyperglycemia, oxidative stress and the subsequent extensive impairment of immune cell functions, a phenomenon responsible for the development of chronic diabetic complications. Propolis, a natural bee product that is extensively used in foods and beverages, significantly benefits human health. Specifically, propolis exerts antioxidant, anti-inflammatory and analgesic effects that may improve diabetic complications. To further elucidate the potential benefits of propolis, the present study investigated the effect of dietary supplementation with propolis on the plasma cytokine profiles, free radical levels, lipid profile and lymphocyte proliferation and chemotaxis in a streptozotocin (STZ)-induced type I diabetic mouse model. METHODS: Thirty male mice were equally distributed into 3 experimental groups: group 1, non-diabetic control mice; group 2, diabetic mice; and group 3, diabetic mice supplemented daily with an ethanol-soluble derivative of propolis (100 mg/kg body weight) for 1 month. RESULTS: First, the induction of diabetes in mice was associated with hyperglycemia and significant decreases in the insulin level and the lymphocyte count. In this context, diabetic mice exhibited severe diabetic complications, as demonstrated by a significant decrease in the levels of IL-2, IL-4 and IL-7, prolonged elevation of the levels of pro-inflammatory cytokines (IL-1ß, IL-6 and TNF-α) and reactive oxygen species (ROS) and altered lipid profiles compared with control non-diabetic mice. Moreover, antigen stimulation of B and T lymphocytes markedly reduced the proliferative capacity and chemotaxis of these cells towards CCL21 and CXCL12 in diabetic mice compared with control mice. Interestingly, compared with diabetes induction alone, treatment of diabetic mice with propolis significantly restored the plasma cytokine and ROS levels and the lipid profile to nearly normal levels. Most importantly, compared with untreated diabetic mice, diabetic mice treated with propolis exhibited significantly enhanced lymphocyte proliferation and chemotaxis towards CCL21 and CXCL12. CONCLUSION: Our findings reveal the potential immuno-modulatory effects of propolis, which acts as a natural antioxidant to enhance the function of immune cells during diabetes.

Modulation of immune cell proliferation and chemotaxis towards CC chemokine ligand (CCL)-21 and CXC chemokine ligand (CXCL)-12 in undenatured whey protein-treated mice.

Whey protein concentrates (WPCs) enhance innate mucosal immunity during early life and have a protective role in some immune disorders. To further elucidate the potential benefits of this protein, the present study investigated the effect of dietary supplementation with WPCs on blood parameters, plasma cytokine profiles, and immune cell proliferation and chemotaxis. A total of 45 male mice were equally distributed into three experimental groups and treated daily for 21 days as follows: group I was a control group that was orally supplemented with distilled water, group II was orally supplemented with undenatured WP (100 mg/kg body weight), and group III was orally supplemented with bovine serum albumin (100 mg/kg body weight). We found that the plasma cytokine levels of interleukin (IL)-1α, IL-1ß, IL-10 and tumor necrosis factor-α and the levels of reactive oxygen species, cholesterol, triglycerides and the lipid profile were significantly decreased in the WP-treated group compared to the control group. In contrast, the levels of IL-2, IL-4, IL-7, IL-8 and glutathione were significantly elevated, and consequently, the ability of peripheral blood mononuclear cells to proliferate in response to stimulation with different antigens was significantly increased in the WP-treated group. Moreover, the in vitro chemotaxis of B, T and bone-marrow-derived dendritic cells toward CC chemokine ligand-21 and CXC chemokine ligand-12 was significantly increased, by twofold, in WP-treated mice compared to the control group. Taken together, our data reveal the benefits of WP supplementation in enhancing immune cell proliferation and migration to the secondary lymphoid organs.

Effects of undenatured whey protein supplementation on CXCL12- and CCL21-mediated B and T cell chemotaxis in diabetic mice.

BACKGROUND: Long and persistent uncontrolled diabetes tends to degenerate the immune system and leads to an increased incidence of infection. Whey proteins (WPs) enhance immunity during early life and have a protective role in some immune disorders. In this study, the effects of camel WP on the chemotaxis of B and T cells to CXCL12 and CCL21 in diabetic mice were investigated. RESULTS: Flow cytometric analysis of the surface expressions of CXCR4 (CXCL12 receptor) and CCR7 (CCL21 receptor) on B and T cells revealed that the surface expressions of CXCR4 and CCR7 were not significantly altered in diabetic and WP-supplemented diabetic mice compared with control mice. Nevertheless, B and T lymphocytes from diabetic mice were found to be in a stunned state, with a marked and significant (P < 0.05) decrease in CXCL12- and CCL21-mediated actin polymerization and subsequently, a marked decrease in their chemotaxis. WP supplementation in the diabetes model was found to significantly increase CXCL12- and CCL21-mediated actin polymerization and chemotaxis in both B and T cells. CONCLUSION: Our data revealed the benefits of WP supplementation in enhancing cytoskeletal rearrangement and chemotaxis in B and T cells, and subsequently improving the immune response in diabetic mice.

Association of a rheumatoid arthritis susceptibility variant at the CCL21 locus with premature mortality in inflammatory polyarthritis patients.

OBJECTIVE: To investigate whether recently identified rheumatoid arthritis (RA) susceptibility loci are also associated with disease severity, specifically all-cause and cardiovascular disease (CVD) mortality, in patients with inflammatory polyarthritis (IP). METHODS: Subjects with recent-onset IP were recruited from the Norfolk Arthritis Register. Seventeen RA susceptibility single-nucleotide polymorphisms (SNPs) were tested using Sequenom MassArray iPLEX chemistry. Vital status was ascertained from central records. The association of SNP allele carriage with mortality risk was assessed using Cox proportional hazards models after adjusting by sex. The mortality risks of those SNP alleles found to be associated were then stratified by baseline anti-citrullinated peptide (anti-CCP) antibody and shared epitope (SE) status. RESULTS: All SNPs were successfully genotyped in 2,324 IP subjects. The presence of 2 copies of the risk allele rs2812378 mapping to the CCL21 gene predicted all-cause mortality (hazard ratio [HR] 1.40, 95% confidence interval [95% CI] 1.04-1.87), whereas risk allele carriage also predicted increased CVD mortality (HR 1.33, 95% CI 1.01-1.75). The highest mortality risks were seen in anti-CCP antibody-positive subjects with 2 copies of the CCL21 risk alleles and 2 copies of the SE (all-cause HR 3.20, 95% CI 1.52-6.72; CVD HR 3.73, 95% CI 1.30-10.72). CONCLUSION: In this large study, we found that carriage of CCL21 risk alleles was associated with premature mortality in IP independently of anti-CCP antibody and SE status. Interestingly, CCL21 expression has been reported in atherosclerotic plaques supporting the thesis that the increased CVD mortality in IP patients may be mediated by shared inflammatory mechanisms.

Developmental and functional studies of the SLC12 gene family members from Drosophila melanogaster.

The electroneutral cation-chloride cotransporter gene family, SLC12, contains nine members in vertebrates. These include seven sodium and/or potassium-coupled chloride transporters and two membrane proteins of unknown function. Although SLC12 family members have been identified in a number of lower species, the functional properties of these proteins are unknown. There are five SLC12 homologues in Drosophila melanogaster, including at least one member on each of the four main branches of the vertebrate phylogenetic tree. We have employed in situ hybridization to study the expression patterns of the Drosophila SLC12 proteins during embryonic development. Our studies indicate that all five members of this family are expressed during early embryogenesis (stages 1-6), but that spatial and temporal expression patterns become more refined as development proceeds. Expression during late embryogenesis was seen predominantly in the ventral nerve cord, salivary gland, gut, and anal pad. In parallel studies, we have carried out transport assays on each of the five Drosophila homologues, expressed as recombinant proteins in the cultured insect cell line High Five. Under our experimental conditions, we found that only one of these proteins, CG4357, transported the potassium congener (86)Rb. Additional experiments established that rubidium transport via CG4357 was saturable (K(m) = 0.29 +/- 0.05 mM), sodium-dependent (half-saturation constant = 53 +/- 11 mM), chloride-dependent (half-saturation constant = 48 +/- 5 mM), and potently inhibited by bumetanide (inhibitor constant = 1.17 +/- 0.08 muM), a specific inhibitor of Na(+)-K(+)-2Cl(-) cotransporters. Taken together, our results provide strong evidence that CG4357 is an insect ortholog of the vertebrate Na(+)-K(+)-2Cl(-) cotransporters.

Effects of bee venom on the maturation of murine dendritic cells stimulated by LPS.

AIM OF STUDY: This study was performed to elicit the effectiveness of bee venom (BV), a traditional immunosuppressive Korean acupuncture agent, on the maturation of dendrtic cells (DCs). MATERIALS AND METHODS: Immature dendritic cells (iDCs) were generated from mouse bone marrow cells with GM-CSF. After 10 days of initial differentiation, DCs were activated with lipopolysaccharides (LPS) for another 48h in the presence or absence of BV. Surface molecule analysis, intracytoplasmic staining of cytokines, FITC-conjugated antigen uptake, and transwell migration assays were conducted with iDCs and activated DCs. RESULTS: Up-regulation of costimulatory molecules, typical of mature DCs (mDCs) was inhibited by addition of BV. Pro-inflammatory cytokines were also found to be reduced with BV treatment in LPS-stimulated DC. A decrease in antigen uptake upon the maturation of DC was reversed in low dose BV treated mDC. In addition, BV treated mDC demonstrated reduced directional migration in response to CCL21, a lymphoid chemokine which directs mDC. CONCLUSIONS: BV may have a therapeutic effect an on abnormally activated immune status, such as autoimmune rheumatoid arthritis, through an immune-modulatory effect on DC.

Pre-clinical characterization of GMP grade CCL21-gene modified dendritic cells for application in a phase I trial in non-small cell lung cancer.

BACKGROUND: Our previous studies have demonstrated that transduction of human dendritic cells (DC) with adenovirus encoding secondary lymphoid chemokine, CCL21, led to secretion of biologically active CCL21 without altering DC phenotype or viability. In addition, intratumoral injections of CCL21-transduced DC into established murine lung tumors resulted in complete regression and protective anti-tumor immunity. These results have provided the rationale to generate a clinical grade adenoviral vector encoding CCL-21 for ex vivo transduction of human DC in order to assess intratumoral administration in late stage human lung cancer. METHODS: In the current study, human monocyte-derived DC were differentiated by exposure to GM-CSF and IL-4 from cryopreserved mononuclear cells obtained from healthy volunteers. Transduction with clinical grade adenoviral vector encoding CCL21 (1167 viral particles per cell) resulted in secretion of CCL21 protein. RESULTS: CCL21 protein production from transduced DC was detected in supernatants (24-72 hours, 3.5-6.7 ng/4-5 x 10(6) cells). DC transduced with the clinical grade adenoviral vector were > 88% viable (n = 16), conserved their phenotype and maintained integral biological activities including dextran uptake, production of immunostimulatory cytokines/chemokines and antigen presentation. Furthermore, supernatant from CCL21-DC induced the chemotaxis of T2 cells in vitro. CONCLUSION: Viable and biologically active clinical grade CCL21 gene-modified DC can be generated from cryopreserved PBMC.

[Impact of secondary lymphoid tissue chemokine on lymphocyte migration and the significance thereof in pathogenesis of ulcerative colitis: experiment with rats].

OBJECTIVE: To explore the impact of secondary lymphoid tissue chemokine (SLC) on lymphocyte migration and the significance thereof in the pathogenesis of ulcerative colitis (UC). METHODS: Sixty SD rats were randomly divided into 3 equal groups: model group undergoing dripping of 40% acetone solution of dinitro-chlorobenzene (DNCB) on the back for 2 weeks and then enema of 6% DNCB acetone solution so as to establish models of UC, and then intravenous injection of normal saline (NS) for 5 days; SLC antibody intervention group undergoing intravenous injection of SLC antibody 15 microg x ml(-1) x kg(-1) immediately after the establishing of model; and control group undergoing enema of NS nly and then intravenous injection of NS for 5 days. Six days after the establishing of model venous blood samples were collected from the portal veins of the 3 groups. Lymphocytes were isolated and cultured. RT-PCR was used to detect the mRNA expression of the SLC receptor CCR7. Boyden chamber system was used to examine the migration ability of the lymphocytes exposed to SLC of 20, 40, 60, 80, and 100 ng/ml respectively. ELISA was used to detect the expression of interleukin (IL)-10 and interferon (IFN)-gamma in the supernatants of the lymphocytes of different groups. RESULTS: RT-PCR showed that the CCR7 mRNA expression level of the model group was (0.792 +/- 0.108), significantly higher than that of the intervention group (0.386 +/- 0.115, P = 0.0429), and the CCR7 mRNA expression levels of these 2 groups were both significantly higher than that of the control group (0.106 +/- 0.029, both P < 0.01). SLC dose-dependently promoted the migration ability of the lymphocytes, but there existed a saturation phenomenon. Exposed to 80 ng/ml SLC the migration level of the lymphocytes of the model group peaked to (85.9 +/- 16.0), 3.7 times as high as that of the control group (20.5 +/- 1.8, P < 0.01), and the migration level of the lymphocytes of the intervention group was 38.2 +/- 6.3, significantly higher than that of the control group too (P < 0.05). SLC enhanced the expression of IFN-gamma of the lymphocytes of the model group, while reduced the IL-10 level, and both effects peaked at the concentration of 80 ng/ml (P = 0.042, P = 0.036). CONCLUSION: SLC promotes the lymphocyte migration and boosts the differentiation of lymphocytes, thus participating in the pathogenesis of UC.

[Abnormal lymphocyte homing and colon lesions in ulcerative colitis: experiment with rats].

OBJECTIVE: To investigate the relationship between abnormal lymphocyte homing and colon lesions in ulcerative colitis. METHODS: 60 Sprague-Dawley rats were randomly divided into 3 equal groups: model group [undergoing enema of dinitrochlorobenzene to establish models of ulcerative colitis and then venous injection of normal saline (NS) once a day for 5 days], lymphocyte homing intervention group [undergoing venous injection of secondary lymphoid-tissue chemokine (SLC) antibody, and then venous injection of NS for 5 days], and control group [undergoing venous injection of NS for 5 days]. On the 6th day blood samples were collected from the portal vein to isolated lymphocytes. Distant colon was dissected to undergo pathological examination of submucosal aggregated lymphatic follicles, ulceration, and inflammation, thus observing the lymphocyte homing situation. Specimens of colon mucosa underwent detection cytokine of interleukin (IL)-2 and IL-6. RT-PCR was used to detect the mRNA expression of SLC gene and the chemokine receptor CCR7. The proportion of CCR7 positive lymphocytes which drainage from colonic vein were measured by flow cytometry (FC). RESULTS: Abnormal lymphocyte homing phenomenon under colonic mucosa was found in the model and intervention groups. The relative grey degree of SLC gene mRNA expression of the model and intervention groups were 0.85 +/- 0.05 and 0.77 +/- 0.14 respectively, both significantly higher than that of the control group (0.31 +/- 0.11, both P < 0.01), however, without significant difference between the 2 former groups. The relative grey degree of CCR7 mRNA expression of the model group was 0.79 +/- 0.11, significantly higher than that of the intervention groups (0.39 +/- 0.12, P = 0.0429), and both were significantly higher than that of the control group (0.11 +/- 0.03, both P < 0.01). FC showed that the proportion of CCR7(+) lymphocytes drainage from colonic vein of the model and the intervention groups were 69% +/- 5% and 77% +/- 10% respectively, both significantly higher than that of the control group (17% +/- 84%, both P < 0.01), however, without significant difference between these 2 former groups (P = 0.0837). CONCLUSION: Abnormal lymphocyte homing is associated with inflammation of the colonic mucosa. Blocking of the lymphocyte homing is effective in reducing the inflammation of colonic mucosa.

Triptolide, a constituent of immunosuppressive Chinese herbal medicine, is a potent suppressor of dendritic-cell maturation and trafficking.

Triptolide (TPT) is a chemically defined, potent immunosuppressive compound isolated from an anti-inflammatory Chinese herbal medicine. TPT has been reported to inhibit autoimmunity, allograft rejection, and graft-versus-host disease (GVHD), and its efficacy was previously attributed to the suppression of T cells. Since dendritic cells (DCs) play a major role in the initiation of T-cell-mediated immunity, we studied the effects of TPT on the phenotype, function, and migration of human monocyte-derived DCs. TPT treatment, over a pharmacologic concentration range, inhibited the lipopolysaccharide (LPS)-induced phenotypic changes, characteristic of mature DCs and the production of interleukin-12p70 (IL-12p70). Consequently, the allostimulatory functions of DCs were impaired by TPT treatment. Furthermore, the calcium mobilization and chemotactic responses of LPS-stimulated DCs to secondary lymphoid tissue chemokine (SLC)/CC chemokine ligand 21 (CCL21) were significantly lower in TPT-treated than untreated DCs, in association with lower chemokine receptor 7 (CCR7) and higher CCR5 expression. Egress of Langerhans cells (LCs) from explanted mouse skin in response to macrophage inflammatory protein-3beta (MIP-3beta)/CCL19 was arrested by TPT. In vivo administration of TPT markedly inhibited hapten (fluorescein isothiocyanate [FITC])-stimulated migration of mouse skin LCs to the draining lymph nodes. These data provide new insight into the mechanism of action of TPT and indicate that the inhibition of maturation and trafficking of DCs by TPT contributes to its immunosuppressive effects.

Role of inducible bronchus associated lymphoid tissue (iBALT) in respiratory immunity.

Bronchus-associated lymphoid tissue (BALT) is occasionally found in the lungs of mice and humans; however, its role in respiratory immunity is unknown. Here we show that mice lacking spleen, lymph nodes and Peyer's patches generate unexpectedly robust primary B- and T-cell responses to influenza, which seem to be initiated at sites of induced BALT (iBALT). Areas of iBALT have distinct B-cell follicles and T-cell areas, and support T and B-cell proliferation. The homeostatic chemokines CXCL13 and CCL21 are expressed independently of TNFalpha and lymphotoxin at sites of iBALT formation. In addition, mice with iBALT, but lacking peripheral lymphoid organs, clear influenza infection and survive higher doses of virus than do normal mice, indicating that immune responses generated in iBALT are not only protective, but potentially less pathologic, than systemic immune responses. Thus, iBALT functions as an inducible secondary lymphoid tissue for respiratory immune responses.

The chemokine CCL21 protects normal marrow progenitors from Ara-C cytotoxicity.

PURPOSE: Chemokines are a family of small proteins that regulate leukocyte infiltration into inflamed tissue and play key roles in the pathogenesis of many diseases. Some chemokines can also reversibly inhibit the proliferation of hematopoietic progenitors. We have previously found that the chemokine CCL21 (Exodus-2/SLC/6Ckine/TCA4) is a potent inhibitor of the proliferation of normal hematopoietic progenitors. In this study we sought to determine whether this inhibition of proliferation could be therapeutically exploited by protecting normal marrow progenitors from the cytotoxicity of the S phase-active chemotherapeutic agent Ara-C. METHODS: Untreated and CCL21-pretreated mice were given doses of Ara-C that are toxic to marrow myeloid progenitors. The recovery of these myeloid progenitors was analyzed by colony formation assays. RESULTS: It was found that pretreatment with small doses of CCL21 prevented the death of normal murine marrow progenitors from the toxic effects of Ara-C. CONCLUSIONS: The chemokine CCL21 may be able to prevent Ara-C myelosuppression during acute leukemia induction chemotherapy, and thereby decrease morbidity and mortality of such therapy, and shorten hospital stays.

Characterization of mouse CCX-CKR, a receptor for the lymphocyte-attracting chemokines TECK/mCCL25, SLC/mCCL21 and MIP-3beta/mCCL19: comparison to human CCX-CKR.

We report here the identification and characterization of murine CCX-CKR, a high affinity receptor for the murine beta-chemokines SLC/mCCL21, MIP-3beta/mCCL19 and TECK/mCCL25. Unlike most other chemokine receptors, CCX-CKR is unable to mediate Ca(2+) fluxes upon ligand binding when expressed in HEK293 cells. Murine CCX-CKR is expressed predominantly in the heart and lung, but is detectable in most organs using RT-PCR. Interestingly, in brain and testis, an alternative mRNA form of CCX-CKR exists with a unique 5' untranslated region (5'UTR) that overlaps with a novel acyl-CoA dehydrogenase (ACD) gene. Analysis of human CCX-CKR shows that the expression profiles and alternative 5'UTR are conserved. However, in man, there are two copies of the CCX-CKR gene, one on chromosome 3 nestled within the ACD homologue, and one on chromosome 6. These genes encode proteins with only one amino acid difference, and their expression is independently regulated. This study identifies murine CCX-CKR, reveals complex regulation of CCX-CKR gene expression in mouse and man, and is suggestive of non-leukocytic targets for MIP-3beta/CCL19, SLC/CCL21 and TECK/CCL25.

Macrophage inflammatory protein-3 beta enhances IL-10 production by activated human peripheral blood monocytes and T cells.

We report that the addition of human macrophage inflammatory protein-3 beta (MIP-3 beta) to cultures of human PBMCs that have been activated with LPS or PHA results in a significant enhancement of IL-10 production. This effect was concentration-dependent, with optimal MIP-3 beta concentrations inducing more than a 5-fold induction of IL-10 from LPS-stimulated PBMCs and a 2- to 3-fold induction of IL-10 from PHA-stimulated PBMCs. In contrast, no significant effect on IL-10 production was observed when 6Ckine, the other reported ligand for human CCR7, or other CC chemokines such as monocyte chemoattractant protein-1, RANTES, MIP-1 alpha, and MIP-1 beta were added to LPS- or PHA-stimulated PBMCs. Similar results were observed using activated purified human peripheral blood monocytes or T cells. Addition of MIP-3 beta to nonactivated PBMCs had no effect on cytokine production. Enhancement of IL-10 production by MIP-3beta correlated with the inhibition of IL-12 p40 and TNF-alpha production by monocytes and with the impairment of IFN-gamma production by T cells, which was reversed by addition of anti-IL-10 Abs to the cultures. The ability of MIP-3 beta to augment IL-10 production correlated with CCR7 mRNA expression and stimulation of intracellular calcium mobilization in both monocytes and T cells. These data indicate that MIP-3 beta acts directly on human monocytes and T cells and suggest that this chemokine is unique among ligands binding to CC receptors due to its ability to modulate inflammatory activity via the enhanced production of the anti-inflammatory cytokine IL-10.