Development of Exhausted Memory Monocytes and Underlying Mechanisms.

Pathogenic inflammation and immuno-suppression are cardinal features of exhausted monocytes increasingly recognized in septic patients and murine models of sepsis. However, underlying mechanisms responsible for the generation of exhausted monocytes have not been addressed. In this report, we examined the generation of exhausted primary murine monocytes through prolonged and repetitive challenges with high dose bacterial endotoxin lipopolysaccharide (LPS). We demonstrated that repetitive LPS challenges skew monocytes into the classically exhausted Ly6Chi population, and deplete the homeostatic non-classical Ly6Clo population, reminiscent of monocyte exhaustion in septic patients. scRNAseq analyses confirmed the expansion of Ly6Chi monocyte cluster, with elevation of pathogenic inflammatory genes previously observed in human septic patients. Furthermore, we identified CD38 as an inflammatory mediator of exhausted monocytes, associated with a drastic depletion of cellular NAD+; elevation of ROS; and compromise of mitochondria respiration, representative of septic monocytes. Mechanistically, we revealed that STAT1 is robustly elevated and sustained in LPS-exhausted monocytes, dependent upon the TRAM adaptor of the TLR4 pathway. TRAM deficient monocytes are largely resistant to LPS-mediated exhaustion, and retain the non-classical homeostatic features. Together, our current study addresses an important yet less-examined area of monocyte exhaustion, by providing phenotypic and mechanistic insights regarding the generation of exhausted monocytes.

Proteomic and molecular evidences of Il1rl2, Ric8a, Krt18 and Hsp90b1 modulation during experimental hepatic fibrosis and pomegranate supplementation.

The inspection of variations in the proteomic aspects conspire the biomarker discovery in diagnostics of peculiar diseases. Recent developments in high-throughput proteomic techniques have provided leverage in the discovery of biomarkers during the etiology of various diseases. We identified potential biomarkers by utilizing proteomics, bioinformatics and gene expression studies. Meticulous assessment of collagen and hydroxyproline levels along with the glycogen and protein carbonyl levels exhibited deterioration in the N' - Nitrosodiethylamine (NDEA) administered rat livers and subsequent salubrious effect of pomegranate juice. The immunohistochemical inspection of iNOS and nitrite estimation indicated the peccant fibrotic alterations. 2D proteome profiling and MALDI-TOF MS/MS furthered the significant biomarkers to be analyzed for the gene ontology by PANTHER, cluster analysis by DAVID and network simulation by STRING 10.0. Several genes found relevant after MALDI analysis were evaluated by real-time PCR (RTPCR). Our data revealed CYP2b15, HSP70, TRFE, HPT, Il1rl2, Ric8a, Krt18, Hsp90b1 and iNOS as novel biomarkers for the mechanism of pomegranate against liver fibrosis. It can be inferred that NDEA-induced liver fibrosis actuates various biological pathways by the identified biomarkers and pomegranate juice modifies them.

Tangeretin attenuates lipopolysaccharide-induced acute lung injury through Notch signaling pathway via suppressing Th17 cell response in mice.

Tangeretin, a polymethoxylated flavonoid is abundant in citrus fruits, which has been reported to inhibit inflammation by inhibiting NF-κB activation and proinflammatory cytokines. Notch blockage inhibits Th17 cells response that are involved in the development of acute lung injury (ALI). This study investigated the protective effects of tangeretin on LPS-induced ALI in mice. Male C57BL/6 mice were treated with phosphate-buffered saline (PBS), lipopolysaccharide (LPS), LPS and tangeretin, or LPS and N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT, a Notch signaling inhibitor), which were harvested at 48 h after challenged by LPS. CD4+ T cells were treated with tangeretin or DAPT and harvested after 72 h. Tangeretin notably attenuated pathological changes and decreased the wet to dry weight ratio of the mouse lungs. The total cell and neutrophil counts, tumor necrosis factor (TNF)-α in bronchoalveolar lavage fluid (BALF), myeloperoxidase activity of lung tissue were markedly reduced by tangeretin. The percentage of CD4+IL-17 + T cells in the lungs and the concentration of interleukin (IL)-17 and IL-22 in BALF were significantly down-regulated by tangeretin. As with the positive control (DAPT), tangeretin inhibited the activity of the Notch signaling pathway accompanied with the down-regulation of acid-related orphan receptor gamma t and IL-23 receptor expression. This study demonstrated that tangeretin protects against LPS-induced ALI by suppressing Th17 response at least partially, through a Notch-dependent mechanism.

Histone Acetylation of Immune Regulatory Genes in Human Placenta in Association with Maternal Intake of Olive Oil and Fish Consumption.

Maternal diet modifies epigenetic programming in offspring, a potentially critical factor in the immune dysregulation of modern societies. We previously found that prenatal fish oil supplementation affects neonatal T-cell histone acetylation of genes implicated in adaptive immunity including PRKCZ, IL13, and TBX21. In this study, we measured H3 and H4 histone acetylation levels by chromatin immunoprecipitation in 173 term placentas collected in the prospective birth cohort, ALADDIN, in which information on lifestyle and diet is thoroughly recorded. In anthroposophic families, regular olive oil usage during pregnancy was associated with increased H3 acetylation at FOXP3 (p = 0.004), IL10RA (p = 0.008), and IL7R (p = 0.007) promoters, which remained significant after adjustment by offspring gender. Furthermore, maternal fish consumption was associated with increased H4 acetylation at the CD14 gene in placentas of female offspring (p = 0.009). In conclusion, prenatal olive oil intake can affect placental histone acetylation in immune regulatory genes, confirming previously observed pro-acetylation effects of olive oil polyphenols. The association with fish consumption may implicate ω-3 polyunsaturated fatty acids present in fish oil. Altered histone acetylation in placentas from mothers who regularly include fish or olive oil in their diets could influence immune priming in the newborn.

Baicalin Ameliorates Imiquimod-Induced Psoriasis-Like Inflammation in Mice.

Baicalin is the main flavonoid from the roots of an important medicinal plant, Scutellaria baicalensis, which shows a variety biological activities. Psoriasis is a chronic immune-mediated inflammatory disease that affects the skin. The unmet need of psoriasis is that many patients do not respond adequately to available clinical treatment. In this study, we found that baicalin showed inhibited dermal inflammation in a murine model of psoriasis via topical application of imiquimod. After a 5-day topical imiquimod application, baicalin or the control vehicle cream was to applied to the lesions of BALB/c mice for a further 4 days. The erythema, scaling, and thickness of the epidermal layer significantly improved in the baicalin-treated mice. The levels of interleukin-17A, interleukin-22, interleukin-23, and tumor necrosis factor in the skin significantly decreased after baicalin treatment. Baicalin also inhibited imiquimod-induced interleukin-17A production in skin draining lymph node cells. The infiltration of γδ T cells into the skin lesions induced by imiquimod was also suppressed after baicalin treatment. These results suggest that baicalin inhibited skin inflammation through the inhibition of the interleukin-17/interleukin-23 axis in a murine model of psoriasis.

Inhibition of Spinal Interlukin-33/ST2 Signaling and Downstream ERK and JNK Pathways in Electroacupuncture Analgesia in Formalin Mice.

Although acupuncture is widely used to manage pain, it remains highly controversial, largely due to the lack of a clear mechanism for its benefits. Here, we investigated the role of IL-33, a novel interleukin (IL)-1 family member, and its receptor ST2 in the analgesic effects of electroacupuncture (EA) on formalin-induced inflammatory pain. The results showed that 1) EA stimulation of ipsilateral Zusanli (ST 36) and Yanglingquan (GB 34) acupoints for 30 min remarkably suppressed the two phases of formalin-induced spontaneous pain; 2) subcutaneous or intrathecal administration of recombinant IL-33 (rIL-33) significantly inhibited the analgesic effect of EA, whereas the ST2 antibody potentiated EA analgesia in formalin mice; 3) EA treatment decreased the up-regulation of IL-33 and ST2 protein following formalin injection; and 4) the suppression of the formalin-induced expression of spinal phosphorylated ERK and JNK induced by EA treatment was significantly attenuated following subcutaneous rIL-33 delivery, and was further decreased by the ST2 antibody. These data suggest that EA alleviates formalin-induced inflammatory pain, at least partially, by inhibiting of spinal IL-33/ST2 signaling and the downstream ERK and JNK pathways.

Crosstalk among IL-23 and DNAX activating protein of 12 kDa-dependent pathways promotes osteoclastogenesis.

IL-23 has been well studied in the context of T cell differentiation; however, its role in the differentiation of myeloid progenitors is less clear. In this paper, we describe a novel role of IL-23 in myeloid cell differentiation. Specifically, we have identified that in human PBMCs, IL-23 induces the expression of MDL-1, a PU.1 transcriptional target during myeloid differentiation, which orchestrates osteoclast differentiation through activation of DNAX activating protein of 12 kDa and its ITAMs. The molecular events that lead to the differentiation of human macrophages to terminally differentiated osteoclasts are dependent on spleen tyrosine kinase and phospholipase Cγ2 phosphorylation for the induction of intracellular calcium flux and the subsequent activation of master regulator osteoclast transcription factor NFATc1. IL-23-elicited osteoclastogenesis is independent of the receptor activator of NF-κB ligand pathway and uses a unique myeloid DNAX activating protein of 12 kDa-associated lectin-1(+)/DNAX activating protein of 12 kDa(+) cell subset. Our data define a novel pathway that is used by IL-23 in myeloid cells and identify a major mechanism for the stimulation of osteoclastogenesis in inflammatory arthritis.

IL-33/ST2 axis promotes mast cell survival via BCLXL.

Mast cells (MC) are potent innate immune cells that accumulate in chronically inflamed tissues. MC express the IL-33 receptor IL-1 receptor-related protein ST2 at high level, and this IL-1 family cytokine both activates MC directly and primes them to respond to other proinflammatory signals. Whether IL-33 and ST2 play a role in MC survival remains to be defined. In skin-derived human MC, we found that IL-33 attenuated MC apoptosis without altering proliferation, an effect mediated principally through the antiapoptotic molecule B-cell lymphoma-X large (BCLXL). Murine MC demonstrated a similar mechanism, dependent entirely on ST2. In line with these observations, St2(-/-) mice exhibited reduced numbers of tissue MC in inflamed arthritic joints, in helminth-infected intestine, and in normal peritoneum. To confirm an MC-intrinsic role for ST2 in vivo, we performed peritoneal transfer of WT and St2(-/-) MC. In St2(-/-) hosts treated with IL-33 and in WT hosts subjected to thioglycollate peritonitis, WT MC displayed a clear survival advantage over coengrafted St2(-/-) MC. IL-33 blockade specifically attenuated this survival advantage, confirming IL-33 as the relevant ST2 ligand mediating MC survival in vivo. Together, these data reveal a cell-intrinsic role for the IL-33/ST2 axis in the regulation of apoptosis in MC, identifying thereby a previously unappreciated pathway supporting expansion of the MC population with inflammation.

Green tea epigallocatechin-3-gallate modulates differentiation of naïve CD4⁺ T cells into specific lineage effector cells.

CD4(+) T helper (Th) subsets Th1, Th9, and Th17 cells are implicated in inducing autoimmunity whereas regulatory T cells (Treg) have a protective effect. We and others have previously shown that epigallocatechin-3-gallate (EGCG) attenuates experimental autoimmune encephalomyelitis (EAE) and alters CD4(+) T cell subpopulations. In this study, we investigated how EGCG impacts differentiation of naïve CD4(+) T cells into different effector lineages and report that EGCG impeded Th1, Th9, and Th17 differentiation and prevented IL-6-induced suppression of Treg development. We further showed that EGCG inhibited T-bet, PU.1, and RORγt, the specific transcription factors for Th1, Th9, and Th17 differentiation, respectively. These effects, in turn, may be mediated by EGCG-induced downregulation of transducers p-STAT1 and p-STAT4 for Th1, and p-STAT3 for Th17. EGCG-induced change in Th17/Treg balance may be mediated by its inhibition of IL-6 signaling because EGCG inhibited soluble IL-6R, membrane gp130, and IL-6-induced phosphorylation of STAT3. This notion was further supported by the in vivo results showing inhibited IL-6 and soluble IL-6R but increased soluble gp130 levels in plasma from EAE mice fed EGCG. Together, our results suggest that EGCG modulates development of CD4(+) T cell lineages through impacting their respective and interactive regulatory networks ultimately leading to an attenuated autoimmune response.

IL-36R ligands are potent regulators of dendritic and T cells.

IL-36α (IL-1F6), IL-36ß (IL-1F8), and IL-36γ (IL-1F9) are members of the IL-1 family of cytokines. These cytokines bind to IL-36R (IL-1Rrp2) and IL-1RAcP, activating similar intracellular signals as IL-1, whereas IL-36Ra (IL-1F5) acts as an IL-36R antagonist (IL-36Ra). In this study, we show that both murine bone marrow-derived dendritic cells (BMDCs) and CD4(+) T lymphocytes constitutively express IL-36R and respond to IL-36α, IL-36ß, and IL-36γ. IL-36 induced the production of proinflammatory cytokines, including IL-12, IL-1ß, IL-6, TNF-α, and IL-23 by BMDCs with a more potent stimulatory effect than that of other IL-1 cytokines. In addition, IL-36ß enhanced the expression of CD80, CD86, and MHC class II by BMDCs. IL-36 also induced the production of IFN-γ, IL-4, and IL-17 by CD4(+) T cells and cultured splenocytes. These stimulatory effects were antagonized by IL-36Ra when used in 100- to 1000-fold molar excess. The immunization of mice with IL-36ß significantly and specifically promoted Th1 responses. Our data thus indicate a critical role of IL-36R ligands in the interface between innate and adaptive immunity, leading to the stimulation of T helper responses.

Contribution of IL-33 to induction and augmentation of experimental allergic conjunctivitis.

IL-33, a member of the IL-1 family of cytokines, is the ligand for ST2 (IL-33Ralpha chain). IL-33 has the capacity to induce T(h)2 cytokine production from T(h)2 cells, mast cells and basophils, indicating that IL-33 has the potential to induce T(h)2 cytokine-mediated allergic inflammation of the eye. Thus, we tested the pathological role of IL-33 in allergic conjunctivitis (AC). As reported elsewhere, animals immunized with ragweed pollen (RW)/alum and boosted with RW/PBS developed AC promptly (within 15 min) and conjunctival eosinophilic inflammation after a delay (within 24 h) in response to eye drop challenge with RW. Furthermore, RW-immunized mice, when topically challenged with both RW and IL-33, developed more striking eosinophilia in their conjunctiva without exacerbation of the clinical AC score. This in vivo IL-33 treatment significantly increased the capacity of T cells in the cervical lymph nodes of RW-immunized mice to produce IL-4, IL-5 and IL-13 upon challenge with anti-CD3 and anti-CD28 antibodies in vitro. Furthermore, the infiltrating cells were largely eosinophils and a small proportion of CD4(+) T cells, both of which express ST2. We also found that even splenic eosinophils express ST2 and show increased expression in response to IL-5, granulocyte-macrophage colony-stimulating factor (GM-CSF) or IL-33. Eosinophils, stimulated with IL-5 and/or GM-CSF, are responsive to IL-33, which induces production of IL-4 and chemokines. Finally, we showed that conjunctival tissues constitutively express biologically active IL-33, suggesting that IL-33 might play a crucial role in the induction and augmentation of AC.

Low level laser therapy (LLLT) decreases pulmonary microvascular leakage, neutrophil influx and IL-1beta levels in airway and lung from rat subjected to LPS-induced inflammation.

BACKGROUND AND OBJECTIVE: Low level laser therapy (LLLT) is a known anti-inflammatory therapy. Herein we studied the effect of LLLT on lung permeability and the IL-1beta level in LPS-induced pulmonary inflammation. STUDY DESIGN/METHODOLOGY: Rats were divided into 12 groups (n = 7 for each group). Lung permeability was measured by quantifying extravasated albumin concentration in lung homogenate, inflammatory cells influx was determined by myeloperoxidase activity, IL-1beta in BAL was determined by ELISA and IL-1beta mRNA expression in trachea was evaluated by RT-PCR. The rats were irradiated on the skin over the upper bronchus at the site of tracheotomy after LPS. RESULTS: LLLT attenuated lung permeability. In addition, there was reduced neutrophil influx, myeloperoxidase activity and both IL-1beta in BAL and IL-1beta mRNA expression in trachea obtained from animals subjected to LPS-induced inflammation. CONCLUSION: LLLT reduced the lung permeability by a mechanism in which the IL-1beta seems to have an important role.

Clinical, immunological, anti-inflammatory and antioxidant roles of zinc.

The essentiality of zinc for humans was recognized only 40 years ago. Zinc deficiency was suspected to occur in Iranian patients with growth retardation, hypogonadism in males, hepato-splenomegaly, rough and dry skin, geophagia and severe iron deficiency anemia. Later we documented zinc deficiency in similar patients in Egypt. The diet of these patients consisted of mainly cereal proteins which contained high phytate and this led to decreased availability of iron and zinc. These patients had severe immune dysfunctions, inasmuch as they died of intercurrent infections by the time they were 25 years of age. In our studies in experimental human model of zinc deficiency, we documented decreased serum testosterone level, oligospermia, severe immune dysfunctions mainly affecting T helper cells, decreased serum thymulin activity hyperammonemia, neuro-sensory disorders and decreased lean body mass. The basic mechanisms of zinc action on immune cells have been reviewed in this paper. Our studies showed that the activation of many zinc dependent enzymes and transcription factors were affected adversely due to zinc deficiency. The gene expression and production of Th1 cytokines were affected adversely due to zinc deficiency. Zinc is also an antioxidant and has anti-inflammatory actions. We have reported decreased plasma zinc, increased plasma oxidative stress markers and increased generation of inflammatory cytokines in the elderly subjects which were corrected by zinc supplementation. In cell culture studies, we have observed that zinc induces A20 which inhibits NF-kappaB activation resulting in decreased generation of inflammatory cytokines.

Immunomodulatory effects of the methanolic extract of Epimedium alpinum in vitro.

The effect of the methanolic extract of root and rhizome of Epimedium alpinum (MEEA) on phenotype and functions of rat lymphocytes in vitro was studied. It has been found that MEEA at lower concentrations (0.1 microg/ml and 1 microg/ml) significantly enhanced proliferation of splenocytes and thymocytes triggered by concanavalin A (Con A), whereas higher concentrations of the extract (50 microg/ml-500 microg/ml) were inhibitory. The stimulatory effect of MEEA on Con A-induced proliferation of splenocytes correlated with the up-regulation of interleukin-2 receptor alpha (IL-2Ralpha) expression. In addition, increased production of IL-2 was observed when a blocking IL-2Ralpha monoclonal antibody (mAb) was added to cell cultures. MEEA-suppressed proliferation of splenocytes was due to the inhibition of IL-2 production, the down-regulation of IL-2Ralpha expression and the induction of apoptosis. Cellular proliferation in the presence of inhibitory concentrations of MEEA higher than 50 microg/ml could not be restored by the addition of exogenous IL-2.

Biological effects of a dietary omega-3 polyunsaturated fatty acids supplementation in cystic fibrosis patients: a randomized, crossover placebo-controlled trial.

BACKGROUND AND AIMS: Various anti-inflammatory therapies, including dietary omega-3 polyunsaturated fatty acids (PUFA) supplementation, have been investigated in cystic fibrosis (CF) patients. To further explore this nutritional approach, biological effects of an omega-3 PUFA oral liquid supplementation were measured in 17 CF patients in a double-blind, randomized, crossover without a washout period and placebo-controlled study. METHODS: CF patients (age: 18+/-9 year; weight: 43+/-13 kg) received a liquid dietary supplementation either enriched or not in omega-3 PUFA (390-1170 mg/day according to patient weight) during two 6-month periods. RESULTS: Increase in eicosapentaenoic acid was observed in neutrophil membrane following omega-3 PUFA dietary supplementation (from 0.7+/-0.6 to 1.6+/-0.6 micromol%, P<0.01). The leukotriene B(4) (LTB(4))/leukotriene B(5) (LTB(5)) ratio was decreased (from 72+/-27 to 24+/-7, P<0.001) in CF patients taking omega-3 PUFA supplements. In contrast, omega-3 PUFA supplementation affected neither internalization of IL-8 receptors following IL-8 exposure, nor IL-8-induced neutrophil chemotaxis. CONCLUSION: Our results show that omega-3 PUFA are incorporated in neutrophil membranes. The subsequent decrease in LTB(4)/LTB(5) ratio suggests that, in such conditions, neutrophils may produce less pro-inflammatory mediators from the acid arachidonic pathway. These data indicate that omega-3 PUFA intake may have anti-inflammatory effect that still need to be assessed by long-term studies following large groups of patients.

Interleukin-13 receptor-directed cytotoxin for malignant glioma therapy: from bench to bedside.

Central nervous system malignant neoplasias, in particular, glioblastoma multiforme (GBM) have defied all current therapeutic modalities. New therapies involving tumor targeting approach are being explored. This approach relies on the identification of unique or over-expressed cell surface receptors or antigens on tumor cells. In that regard, we have identified receptor for an immune regulatory cytokine, interleukin-13 (IL-13), which is over-expressed on human malignant glioma cell lines and primary tumor cell cultures. To target IL-13 receptors (IL-13R) for cancer therapy, we have developed a recombinant fusion protein composed of IL-13 and a mutated form of Pseudomonas exotoxin (IL13-PE38QQR or IL-13 cytotoxin). The IL-13 cytotoxin was found to be highly selective and potent in killing human GBM cells in vitro while normal cells including immune cells, endothelial cells and normal brain cells were generally spared the cytotoxic effect of IL-13 cytotoxin. This is because these cells either expressed none or expressed low levels of IL-13R. Consistent with in vitro cytotoxic activity, IL-13 cytotoxin mediated remarkable anti-tumor activity to human glioma in animal xenograft models. The direct injection of IL-13 cytotoxin into subcutaneous human GBM tumors grown in nude mice produced complete and durable regression of established tumors. Intravenous and intraperitoneal administration of IL-13 cytotoxin also reduced tumor burden significantly with fewer complete responders. All animals tolerated therapy well with minimal toxicity to vital organs. Pre-clinical safety and toxicity studies were performed in mice, rats and monkeys. Systemic administration of IL-13 cytotoxin appeared to be well tolerated at high doses (up to 50 microg/kg). Intrabrain parenchyma administration of IL-13 cytotoxin at doses up to 100 microg/ml was very well tolerated without any evidence of gross or microscopic necrosis, whereas at 500 microg/ml dose, localized necrosis was observed in normal rat brain. Based on these encouraging pre-clinical studies, three Phase I/II clinical trials in adults with malignant glioma have been initiated. The first clinical trial involves convection-enhanced delivery (CED) of IL-13 cytotoxin into recurrent malignant glioma. This route of IL-13 cytotoxin administration appears to be fairly well tolerated with no neurotoxicity. The second clinical trial involves infusion of IL-13 cytotoxin by CED following tumor resection. The initial stage of the second study assessed histologic effect of drug administered prior to resection. In third one, IL-13 cytotoxin is infused by CED followed by tumor resection. All three clinical trials are currently ongoing.

In vitro allergen-induced mRNA expression of signaling lymphocytic activation molecule by PBMC of patients with allergic rhinitis is increased during specific pollen immunotherapy.

BACKGROUND: Specific immunotherapy (SIT) acts by inducing a shift from T(H)2 to T(H)1 cell response on mucous membranes, reducing allergic inflammation. New genes expressed primarily in T(H)1-type cells have been found. Of these genes, signaling lymphocytic activation molecule (SLAM) promotes T-cell proliferation and IFN-gamma production. Nothing is known about its role in T(H)2-T(H)1 switch during SIT. OBJECTIVE: We sought to analyze the mRNA expression of SLAM and other T(H)1-associated genes, interleukin-12 receptor beta2 (IL-12Rbeta2) and T-box expressed in T cells (T-bet), and compare them with the clinical outcome of the therapy. METHODS: PBMC from 30 patients allergic to pollen undergoing SIT were collected during the therapy. Control PBMC were collected from 10 patients with allergic rhinitis not participating in SIT and from 10 nonallergic subjects. Cells were stimulated in vitro with pollen allergen extracts. SLAM, IL-12Rbeta2, and T-bet mRNA expressions were studied by real-time quantitative RT-PCR technique (Taqman). Symptom scoring and medication scoring were registered before commencement of SIT and after 1 year of the therapy. RESULTS: Before the treatment, in vitro allergen-induced SLAM mRNA expression in PBMC was significantly lower in the patients with allergic rhinitis than in the healthy control subjects. After 1 year of the treatment, SLAM mRNA expression was increased in the patients undergoing SIT and was associated with IFN-gamma mRNA expression and inversely associated with the symptom improvement. At the maintenance dose, an increase in SLAM mRNA expression was associated with the clinical symptom improvement at 1 year. No changes were seen in IL-12Rbeta(2) or T-bet mRNA expressions. CONCLUSIONS: SLAM mRNA expression in PBMC is modulated during the course of SIT, and an early and transient increase of SLAM mRNA expression is associated with clinical symptom improvement.

Changes in lymphokine receptor expression and fatty acid composition of phospholipids and triacylglycerols in rat adipocytes associated with lymph nodes following a transient immune challenge.

Single-photon counting fluorimetry was used to record the time course of the expression of interleukin-10 receptors labelled with fluorescent antibodies on the surface of adipocytes over 24h, following an immune challenge to the rat popliteal lymph node. Homologous perinodal and remote-from-node samples from the stimulated and unstimulated popliteal depots were compared in rats fed on plain chow and chow supplemented with 10% w/w suet, fish or vegetable oils. Receptor expression was maximal 6 h after stimulation, and returned to baseline after 24 h, and was similar in the stimulated and unstimulated depots. Fewer receptors were elicited in tissues from rats fed lipid-supplemented diets compared with the control diet, with fewest of all following the fish oil diet. These data suggest that interleukin-10 is involved in local interactions between perinodal adipocytes and lymph node lymphoid cells. Both triacylglycerols and phospholipids contained more polyunsaturates and fewer saturates in perinodal adipose tissue than in samples from sites not associated with lymphoid tissue. These data are consistent with paracrine interactions between perinodal adipocytes and activated lymphoid cells.

Activated STAT4 has an essential role in Th1 differentiation and proliferation that is independent of its role in the maintenance of IL-12R beta 2 chain expression and signaling.

In this study we demonstrated that CD4(+) T cells from STAT4(-/-) mice exhibit reduced IL-12R expression and poor IL-12R signaling function. This raised the question of whether activated STAT4 participates in Th1 cell development mainly through its effects on IL-12 signaling. In a first approach to this question we determined the capacity of CD4(+) T cells from STAT4(-/-) bearing an IL-12Rbeta2 chain transgene (and thus capable of normal IL-12R expression and signaling) to undergo Th1 differentiation when stimulated by Con A and APCs. We found that such cells were still unable to exhibit IL-12-mediated IFN-gamma production. In a second approach to this question, we created Th2 cell lines (D10 cells) transfected with STAT4-expressing plasmids with various tyrosine-->phenylalanine mutations and CD4(+) T cell lines from IL-12beta2(-/-) mice infected with retroviruses expressing similarly STAT4 mutations that nevertheless express surface IL-12Rbeta2 chains. We then showed that constructs that were unable to support STAT4 tyrosine phosphorylation (in D10 cells) as a result of mutation were also incapable of supporting IL-12-induced IFN-gamma production (in IL-12Rbeta2(-/-) cells). Thus, by two complementary approaches we demonstrated that activated STAT4 has an essential downstream role in Th1 cell differentiation that is independent of its role in the support of IL-12Rbeta2 chain signaling. This implies that STAT4 is an essential element in the early events of Th1 differentiation.