Many survivors of breast cancer show significant cognitive impairments, including memory deficits. Inflammation induced by chemotherapy may contribute to hippocampal changes that underlie these deficits. In this cross-sectional study, we measured bilateral hippocampal volumes from high-resolution magnetic resonance images in 42 chemotherapy-treated breast cancer survivors and <em>35</em> healthy female controls. Patients with breast cancer were, on average, 4.8 ± 3.4 years off-therapy. In a subset of these participants (20 breast cancer, 23 controls), we quantified serum cytokine levels. Left hippocampal volumes and memory performance were significantly reduced and <em>interleukin</em>-6 (IL-6) and tumor necrosis factor-alpha (TNFα) concentrations were significantly elevated in the breast cancer group compared to controls. In the breast cancer group, lower left hippocampal volume was associated with higher levels of TNFα and lower levels of IL-6 with a significant interaction between these two cytokines suggesting a potential modulatory effect of IL-6 on TNFα. Verbal memory performance was associated with cytokine levels and left hippocampal volume in both groups. These findings provide evidence of altered hippocampal volume and verbal memory difficulties following breast cancer chemotherapy that may be mediated by TNFα and IL-6.

OBJECTIVE

Inflammation and infection play a major role in preterm birth. The purpose of this study was to (i) determine the prevalence and clinical significance of sterile intra-amniotic inflammation and (ii) examine the relationship between amniotic fluid (AF) concentrations of high mobility group box-1 (HMGB1) and the interval from amniocentesis to delivery in patients with sterile intra-amniotic inflammation.

METHODS

AF samples obtained from 135 women with preterm labor and intact membranes were analyzed using cultivation techniques as well as broad-range PCR and mass spectrometry (PCR/ESI-MS). Sterile intra-amniotic inflammation was defined when patients with negative AF cultures and without evidence of microbial footprints had intra-amniotic inflammation (AF interleukin-6 ≥ 2.6 ng/mL).

RESULTS

(i) The frequency of sterile intra-amniotic inflammation was significantly greater than that of microbial-associated intra-amniotic inflammation [26% (35/135) versus 11% (15/135); (P = 0.005)], (ii) patients with sterile intra-amniotic inflammation delivered at comparable gestational ages had similar rates of acute placental inflammation and adverse neonatal outcomes as patients with microbial-associated intra-amniotic inflammation, and (iii) patients with sterile intra-amniotic inflammation and high AF concentrations of HMGB1 (≥8.55 ng/mL) delivered earlier than those with low AF concentrations of HMGB1 (P = 0.02).

CONCLUSIONS

(i) Sterile intra-amniotic inflammation is more frequent than microbial-associated intra-amniotic inflammation, and (ii) we propose that danger signals participate in sterile intra-amniotic inflammation in the setting of preterm labor.

Inflammation is a critical factor for development of hypoxic-ischemic (HI) brain injury. <em>Interleukin</em>-18 (IL-18) is a proinflammatory cytokine expressed in microglia and processed by caspase-1. Our aim was to characterize the expression of IL-18 and its receptor in relation to caspase-1 and IL-1beta after HI and to evaluate to what extent IL-18 contributes to HI brain injury. Seven-day-old rats were subjected to HI, and brain tissue was sampled at different time points (3 hr to 14 d) after insult. The mRNA for IL-18 and caspase-1 were analyzed with reverse transcriptase PCR, protein was analyzed by Western blot (IL-18, caspase-1) or ELISA (IL-1beta), and the regional distribution was assessed by immunohistochemistry. HI was also induced in C57BL/6 mice, and brain injury in IL-18-deficient animals was compared with that in wild-type animals. The expression of mRNA/protein for caspase-1 and IL-18 in brain homogenates increased progressively at 12 hr to 14 d after HI, whereas IL-1beta peaked at 8 hr. A widespread expression of caspase-1 and IL-18 protein in microglia was found in the HI hemisphere. The IL-18 receptor was expressed on neurons of the cerebral cortex and thalamus. IL-1beta was primarily found in microglia in the habenular nucleus of the thalamus. The infarct volume was reduced by 21% (p = 0.01), and the neuropathology score was significantly decreased in the cerebral cortex (-<em>35</em>%), hippocampus (-22%), striatum (-18%), and thalamus (-17%) in mice with IL-18 deficiency compared with wild-type mice. In conclusion, we found that IL-18 expression in microglia was markedly increased after HI and that IL-18 appears to be important for the development of HI brain injury.

Advances in cytokine biology have helped us understand the complex communication that takes place between antigen-presenting cells and cells of the adaptive immune system, such as T cells, which collectively mediate an appropriate immune response to a plethora of pathogens while maintaining tolerance to self-antigens. The <em>interleukin</em>-12 (IL-12) cytokine family remains one of the most important and includes IL-12, IL-23, IL-27, and the recently identified IL-<em>35</em>. All four are heterodimeric cytokines, composed of an alpha chain (p19, p28, or p<em>35</em>) and a beta chain (p40 or Ebi3), and signal through unique pairings of five receptor chains (IL-12Rbeta1, IL-12Rbeta2, IL-23R, gp130, and WSX-1). Despite the interrelationship between the cytokines themselves and their receptors, their source, activity, and kinetics of expression are quite different. Studies using genetically deficient mice have greatly enhanced our understanding of the biology of these cytokines. However, interpretation of these data has been complicated by the recent realization that p40(-/-), p<em>35</em>(-/-), and Ebi3(-/-) mice all lack more than one cytokine (IL-12/IL-23, IL-12/IL-<em>35</em>, and IL-27/IL-<em>35</em>, respectively). In this review, we compare and contrast the biology of this expanded IL-12 family and re-evaluate data derived from the analysis of these dual cytokine-deficient mice. We also discuss how the opposing characteristics of the IL-12 family siblings may help to promote a balanced immune response.

OBJECTIVE

To compare the singular and combined effects of tumor necrosis factor alpha (TNFalpha), interleukin-1beta (IL-1beta), and IL-17 on messenger RNA (mRNA) expression, translation, and secretion of IL-6, IL-8, and IL-1beta in fibroblasts.

METHODS

Fibroblasts were stimulated with the relevant cytokine(s), pulse labeled with 35S-methionine, and the newly synthesized proteins were immunoprecipitated and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Gene expression was determined by Northern blot analysis. Secreted proteins were detected by enzyme-linked immunosorbent assay (ELISA).

RESULTS

IL-17 alone was a weaker stimulator of the transcription, translation, and secretion of other interleukins than was TNFalpha or IL-1beta. IL-17 (10 ng/ml) stimulated the expression of IL-6 mRNA by 1.3-fold, while TNFalpha (1 ng/ml) increased it by 3.7-fold, and IL-1beta (0.1 ng/ml) increased it by >30-fold. Unlike TNFalpha and IL-1beta, IL-17 hardly affected the expression of IL-8 and IL-1beta mRNA. Translation of IL-6 was 6.2 times greater with IL-17, but TNFalpha and IL-1beta stimulated it 28.9- and 174-fold, respectively. ELISA-measured secretion of IL-6 and IL-8 increased by 6.7 and 5.8 times, respectively, with IL-17, compared with 52 and 269 times with TNFalpha stimulation and 1,356 and 1,084 times with IL-1beta stimulation. Yet, when IL-17 was combined with other cytokines, these activities were stimulated much beyond the sum of the individual effects. The combination of IL-17 and TNFalpha induced the expression of IL-6 or IL-1beta mRNA 7 times more than their additive stimulation, and that of IL-8 mRNA 3.8 times more. Likewise, the secretion of IL-6 and IL-8 was 20 times and 5 times higher, respectively, than expected. This synergism started after 4 hours of combined treatment, and decayed after 24-48 hours regardless of cytokine presence. It could be blocked with anti-IL-17 but not with anti-IL-1.

CONCLUSIONS

Our findings suggest that the primary role of IL-17 is to synergize with TNFalpha and to fine-tune the inflammation process. Therefore, IL-17 may be a potential target for therapeutic intervention.

Circulating peripheral blood polymorphonuclear neutrophils (PMNs) have long been considered terminally differentiated cells that do not synthesize or secrete protein. However, work of others and ourselves has shown that PMNs can secrete the cytokine <em>interleukin</em> 1. In the present study we investigated whether circulating PMNs are capable of synthesizing and secreting another cytokine, tumor necrosis factor alpha (TNF-alpha). Highly purified (greater than 99% granulocytes) PMNs were isolated from normal human volunteer blood and cultured with or without bacterial lipopolysaccharide (LPS) for up to 24 hr. Cell culture supernatants were collected and tested for TNF-alpha, and total RNA was isolated from cells at various times after stimulation and assessed for TNF-alpha mRNA by Northern blot techniques. The results showed that message for TNF-alpha was produced after 60 min of in vitro stimulation with LPS and was maximal at about 4 hr. TNF-alpha was secreted into the supernatant of unstimulated PMNs from two different donors during 24 hr of culture (<em>35</em>-50 pg/ml), but significantly more (160-190 pg/ml) was secreted by PMNs when stimulated with LPS. PMNs from six other normal volunteers showed significant LPS-stimulated secretion of TNF at 60-180 min of culture. The secreted product also had biological activity against the TNF-sensitive L-M cell line, confirming that PMNs can make and secrete immunologically and biologically active TNF. Since it is also possible for monocytes to synthesize and secrete TNF, the amount of TNF secreted by a monocyte population equal to 20% of the PMNs cultured was measured. The results showed that monocytes at a concentration 20 times that potentially contaminating the PMN populations cultured could not produce as much TNF (unstimulated, 26-65 pg/ml; stimulated, 32-87 pg/ml). The PMN must now be considered a cell capable of altering the acute inflammatory response and modulating the immune response through the synthesis and release of cytokines.

BACKGROUND

A human immunodeficiency virus (HIV) vaccine that limits disease and transmission is urgently needed. This clinical trial evaluated the safety and immunogenicity of an HIV vaccine that combines a plasmid-DNA priming vaccine and a modified vaccinia virus Ankara (MVA) boosting vaccine.

METHODS

Forty healthy volunteers were injected with DNA plasmids containing gp160 of HIV-1 subtypes A, B, and C; rev B; p17/p24 gag A and B, and RTmut B by use of a needle-free injection system. The vaccine was administered intradermally or intramuscularly, with or without recombinant granulocyte macrophage colony-stimulating factor, and boosted with a heterologous MVA containing env, gag, and pol of CRF01A_E. Immune responses were monitored with HIV-specific interferon (IFN)-gamma and interleukin (IL)-2 ELISpot and lymphoproliferative assays (LPAs).

RESULTS

Vaccine-related adverse events were mild and tolerable. After receipt of the DNA priming vaccine, 11 (30%) of 37 vaccinees had HIV-specific IFN-gamma responses. After receipt of the MVA boosting vaccine, ELISpot assays showed that 34 (92%) of 37 vaccinees had HIV-specific IFN-gamma responses, 32 (86%) to Gag and 24 (65%) to Env. IFN-gamma production was detected in both the CD8(+) T cell compartment (5 of 9 selected vaccinees) and the CD4(+) T cell compartment (9 of 9). ELISpot results showed that 25 (68%) of 37 vaccinees had a positive IL-2 response and 35 (92%) of 38 had a positive LPA response. Of 38 subjects, a total of 37 (97%) were responders. One milligram of HIV-1 DNA administered intradermally was as effective as 4 mg administered intramuscularly in priming for the MVA boosting vaccine.

CONCLUSIONS

This HIV-DNA priming-MVA boosting approach is safe and highly immunogenic.

BACKGROUND

International Standard Randomised Controlled Trial number: ISRCTN32604572 .

OBJECTIVE

The aim of the study was to assess the incidence and clinical implications of increased plasma angiotensin II despite chronic ACE inhibitor therapy in patients with heart failure.

RESULTS

The studied population consisted of 70 patients (mean age 59+/-9 years). Plasma renin activity and plasma concentration of aldosterone, norepinephrine, atrial natriuretic peptide, angiotensin II, tumour necrosis factor, <em>interleukin</em>-6 and <em>interleukin</em>-1B were assessed at 6 months of ACE inhibitor therapy. Mean left ventricular ejection fraction was 24+/-5% and the end-systolic and end-diastolic diameters were 59+/-9 and 71+/-8 mm, respectively. Despite chronic enalapril or captopril therapy, <em>35</em> patients (50%) had increased plasma angiotensin II (median 33 pg. ml(-1), range 17-84), while it was in the normal range in the remaining <em>35</em> patients (median 10 pg. ml(-1), range 5-15). Plasma renin activity (P=0.005), <em>interleukin</em>-6 (P=0.004), New York Heart Association functional class III-IV (P=0. 006), furosemide dose (P=0.01), lack of beta-blocker therapy (P=0. 04) and norepinephrine (P=0.04) were univariately associated with increased angiotensin II. Multivariate regression analysis identified the plasma renin activity (0.0004), norepinephrine (0.02) and <em>interleukin</em>-6 (0.03) as independent predictors of plasma angiotensin II. During follow-up (<em>35</em>+/-29 months), nine (12.8%) patients died and 13 had new heart failure episodes. Increased plasma angiotensin II, despite ACE inhibitor therapy, was a significant predictor of death or heart failure according to the Kaplan-Meier survival method by log rank test (P=0.002).

CONCLUSIONS

Fifty per cent of patients with heart failure, ha increased plasma angiotension II despite chronic ACE inhibitor therapy. These patients had higher neurohormonal activation and poor prognosis.

It has been suggested that neuroimmunologic mechanisms may be involved in the development and maintenance of neuropathic pain. To further address this concept, the immunoreactive spinal expression of the pro-inflammatory cytokine, <em>interleukin</em>-6 (IL-6), was determined in the mononeuropathy model in the rat, sciatic cryoneurolysis (SCN). This well-established animal model expresses behaviors suggestive of neuropathic pain in humans. Immunohistochemical localization in the spinal cord was determined at 3, 7, 14, 21, <em>35</em>, and 120 days after SCN (n = 6 per time point). Immunoreactive IL-6 increased incrementally in the substantia gelatinosa and motoneurons over time following SCN as compared with normal rats. In an additional study, recombinant human IL-6 was administered intrathecally to normal and previously SCN-lesioned rats. Intrathecal IL-6 produced touch-evoked allodynia (increased sensitivity to a nonnoxious stimulus) in normal rats and thermal hyperalgesia (increased sensitivity to a noxious stimulus) in previously lesioned SCN rats. These results provide evidence that IL-6 may be involved in the cascade of events leading to the development and maintenance of behaviors suggestive of neuropathic pain following peripheral nerve injury.

The heparin-containing mast cells that reside in the connective tissue of the mouse, but not the chondroitin sulfate-containing mast cells in the gastrointestinal mucosa, stain with safranin when exposed to alcian blue/safranin. Mouse bone marrow-derived mast cells (BMMC), the probable in vitro counterparts of in vivo mucosal mast cells, were cultured for 14 days with mouse skin-derived 3T3 fibroblasts in RPMI 1640 medium containing 10% fetal calf serum and 50% WEHI-3 conditioned medium. Although the BMMC adhered to the fibroblast monolayer, they continued to divide, probably due to the presence of <em>interleukin</em> 3 in the conditioned medium. The mast cells remained viable throughout the period of coculture, since they failed to release lactate dehydrogenase and because they increased their histamine content approximately 15-fold. After 12-14 days of coculture, greater than 50% of the BMMC changed histochemically to become safranin+; 30-40% of the <em>35S</em>-labeled glycosaminoglycans on the proteoglycans synthesized by these cocultured mast cells were heparin, whereas heparin was not detected in the initial BMMC. In the absence of WEHI-3 conditioned medium, BMMC adhered to the fibroblast monolayer, and after 8 days of coculture, the number of mast cells did not change and their histamine content remained the same. However, these mast cells also became safranin+ and synthesized 40% heparin glycosaminoglycans. Thus, coculture of BMMC with fibroblasts induces a phenotypic change so that the resulting mast cells stain safranin+ and synthesize heparin proteoglycans, whereas the presence of WEHI-3 conditioned medium stimulates proliferation and an increase in histamine content.

Precursors of activated killer (AK) cells cytotoxic for human noncultured metastatic melanoma and colon carcinoma were characterized. These cells required 3 days incubation with recombinant <em>interleukin</em> 2 (rIL 2) and DNA synthesis for the induction of AK activity. Both negative and positive cell purification methods were used to identify the subpopulation of cells containing AK precursors. By complement-mediated cell depletion studies, AK precursors were largely present in the Leu-11+ fraction, and to a much lesser extent in the Leu-7+ and Leu-2a+ fractions; they were absent in Leu-3a+ and Leu-4+ cells. Lymphocyte subpopulations were then purified with a cell sorter to positively select for the subset containing AK precursors. Leu-11+ cells had the highest level of AK activity and proliferative response when cultured for 3 days with rIL 2 as well as the highest level of NK activity before culture. Leu-7+ cells had neither AK activity nor a proliferative response when cultured with rIL 2, although they still possessed high NK activity. The same levels of AK and NK activity were found in Leu-2a+ and Leu-2a- fractions, but both activities were absent among Leu-4+ and Leu-3a+ cells. Further fractionation with a two-step sorting technique showed that the highest AK activity resided in the Leu-7-Leu-11+ cell fraction. Morphologically, this subfraction was granular lymphocytes. Titration experiments or rIL 2-responsive cells showed that the number of cells required to achieve a comparable level of rIL 2 proliferative response were as follows: <em>35</em> X 10(3) cells from unseparated PBL, 10 X 10(3) cells from Leu-11+ cells, 3.3 X 10(3) from Leu-7-Leu-11+ cells, and 640 X 10(3) cells from Leu-7+ cells. These results indicate that the lymphocyte subpopulation that proliferates in the presence of rIL 2 and then develops AK activity was a subpopulation of Leu-11+ granular lymphocytes, which also possessed the highest NK activity. These Leu-11+ cells lacked the antigens defined by the Leu-7, Leu-3a, or Leu-4 antibodies. Although Leu-7+ cells did not respond to rIL 2 by themselves, they may play a role in the induction of AK activity.

The copper chelator tetraethylenepentamine (TEPA; StemEx) was shown to attenuate the differentiation of ex vivo cultured hematopoietic cells resulting in preferential expansion of early progenitors. A phase I/II trial was performed to test the feasibility and safety of transplantation of CD133+ cord blood (CB) hematopoietic progenitors cultured in media containing stem cell factor, FLT-3 ligand, <em>interleukin</em>-6, thrombopoietin and TEPA. Ten patients with advanced hematological malignancies were transplanted with a CB unit originally frozen in two fractions. The smaller fraction was cultured ex vivo for 21 days and transplanted 24 h after infusion of the larger unmanipulated fraction. All but two units contained <2 x 10(7) total nucleated cells (TNCs) per kilogram pre-expansion. All donor-recipient pairs were mismatched for one or two HLA loci. Nine patients were beyond first remission; median age and weight were 21 years and 68.5 kg. The average TNCs fold expansion was 219 (range, 2-620). Mean increase of CD34+ cell count was 6 (over the CD34+ cell content in the entire unit). Despite the low TNCs per kilogram infused (median=1.8 x 10(7)/kg), nine patients engrafted. Median time to neutrophil and platelet engraftment was 30 (range, 16-46) and 48 (range, <em>35</em>-105) days. There were no cases of grades 3-4 acute graft-versus-host disease (GVHD) and 100-day survival was 90%. This strategy is feasible.

OBJECTIVE

Obese patients demonstrate a variety of biochemical, metabolic, and pulmonary abnormalities. Inflammatory mediators such as tumor necrosis factor-alpha and interleukin-6 (IL-6) may have a direct effect on glucose and lipid metabolism. Hypoxemia in itself induces release of IL-6. The aim of this study was to examine the relationship between IL-6 levels in healthy volunteers (control group) and three different groups of obese patients: patients without obstructive sleep apnea syndrome (OSAS), patients with OSAS, and patients with obesity hypoventilation syndrome (OHS) (daytime baseline oxygen saturation of <93%).

METHODS

We measured serum IL-6 levels in 25 obese patients (body mass index of >35 kg/m2) and 12 healthy women.

RESULTS

The results demonstrate statistically significant differences in serum IL-6 levels between the control group (1.28 +/- 0.85 pg/mL) and obese patients without OSAS (7.69 +/- 5.06 pg/mL, p < 0.05) and with OSAS (5.58 +/- 0.37 pg/mL, p < 0.0005). In the patients with OHS, IL-6 concentrations were highest (43.13 +/- 24.27 pg/mL).

CONCLUSIONS

We conclude that serum IL-6 is increased in obese patients. The highest IL-6 levels were found in the patients with OHS.

It remains unknown whether newly identified anti-inflammatory/immunosuppressive cytokine <em>interleukin</em>-<em>35</em> (IL-<em>35</em>) is different from other anti-inflammatory cytokines such as IL-10 and transforming growth factor (TGF)-β in terms of inhibition of inflammation initiation and suppression of full-blown inflammation. Using experimental database mining and statistical analysis methods we developed, we examined the tissue expression profiles and regulatory mechanisms of IL-<em>35</em> in comparison to other anti-inflammatory cytokines. Our results suggest that in contrast to TGF-β, IL-<em>35</em> is not constitutively expressed in human tissues but it is inducible in response to inflammatory stimuli. We also provide structural evidence that AU-rich element (ARE) binding proteins and microRNAs target IL-<em>35</em> subunit transcripts, by which IL-<em>35</em> may achieve non-constitutive expression status. Furthermore, we propose a new system to categorize anti-inflammatory cytokines into two groups: (1) the house-keeping cytokines, such as TGF-β, inhibit the initiation of inflammation whereas (2) the responsive cytokines including IL-<em>35</em> suppress inflammation in full-blown stage. Our in-depth analyses of molecular events that regulate the production of IL-<em>35</em> as well as the new categorization system of anti-inflammatory cytokines are important for the design of new strategies of immune therapies.

The suppressor of cytokine signaling (SOCS) family of proteins has been implicated in the negative regulation of several cytokine pathways, particularly the receptor-associated tyrosine kinase/signal transducer and activator of transcription (Jak/STAT) pathways of transcriptional activation. SOCS-1 (also known as JAB and SSI-1) inhibits signaling by many cytokines. Because of the previously observed hypermethylation-associated inactivation of SOCS-1 in hepatocellular carcinoma and the critical role of <em>interleukin</em>-6 (IL-6) as a survival factor in multiple myeloma (MM), we examined CpG island methylation of the SOCS-1 gene in MM cell lines and primary MM samples. Aberrant SOCS-1 methylation was found in the IL-6-dependent MM cell lines U266 and XG1, which correlated with transcriptional silencing. Treatment of these cell lines with the demethylating agent 5-aza-2'-deoxycytidine (DAC) up-regulated SOCS-1 expression. Methylation-associated inactivation of SOCS-1 in hematopoietic cell lines correlated with greater sensitivity to the chemical JAK inhibitor AG490. Using methylation-specific polymerase chain reaction (MSP), we found that SOCS-1 is hypermethylated in 62.9% (23/<em>35</em>) of MM patient samples. In contrast, methylation analysis of malignant lymphomas of various histologies revealed SOCS-1 hypermethylation in only 3.2% (2/62), and there was no methylation of SOCS-1 in normal peripheral blood leukocytes or bone marrow cells. We conclude that SOCS-1 is frequently inactivated by hypermethylation in MM patients. Silencing of the SOCS-1 gene may impair negative regulation of the Jak/STAT pathway and therefore result in greater responsiveness to cytokines, thus supporting survival and expansion of MM cells.

OBJECTIVE

Diabetic retinopathy is shown to share many similarities with chronic inflammatory disease, and in diabetes, accelerated apoptosis of retinal capillary cells is evident before histopathology can be seen. The purpose of this study was to examine the effect of interleukin (IL)-1beta on capillary cell apoptosis in rat retina and to determine the effect of antioxidants on diabetes-induced changes in retinal IL-1beta.

METHODS

The effect of injection of IL-1beta into the vitreous (5 ng/5 microL) of normal rats on capillary cell apoptosis (detected by terminal transferase dUTP nick-end labeling [TUNEL]) and formation of acellular capillaries was investigated in the trypsin digested retinal microvessels. The levels of IL-1beta were quantified (by ELISA and Western blot) in the retina of rats diabetic for 2 months, and the effect of administration of antioxidants on diabetes-induced changes in retinal IL-1beta was determined.

RESULTS

The number of TUNEL-positive capillary cells in the retinal microvessels obtained from normal rats that received intravitreal injection of IL-1beta was increased by more than threefold and that of acellular capillaries by more than twofold, compared with the microvessels obtained from rats that received an intravitreal injection of PBS (5 microL) or BSA (5 ng/5 microL). IL-1beta also increased the levels of 8-hydroxy-2'-deoxyguanosine (an indicator of oxidative stress) and nitric oxide by more than 40% and activated NF-kappaB by 35% to 55%. Two months of diabetes in rats increased retinal IL-1beta levels by more than twofold, and antioxidants inhibited such increases.

CONCLUSIONS

IL-1beta, by activation of NF-kappaB and an increase in oxidative stress, plays an important role in the retinal microvascular disease that is characteristic of diabetic retinopathy. Antioxidants inhibit diabetes-induced increases in retinal IL-1beta. These studies offer a possible rationale to test IL-1beta-targeted therapies to inhibit the development of retinopathy in diabetes.

OBJECTIVE

The purpose of this study was to examine the influence of a 12-wk exercise training program on inflammatory cytokine and C-reactive protein (CRP) concentrations. A secondary purpose was to determine whether training-induced changes in cytokines and CRP were influenced by age.

METHODS

Twenty-nine younger (18-<em>35</em> yr) and 31 older (65-85 yr) subjects were assigned to young physically active (YPA, N = 15; 25 +/- 5 yr), young physically inactive (YPI, N= 14; 25 +/- 4.7 yr), old physically active (OPA, N = 14; 71 +/- 4 yr), or old physically inactive (OPI, N = 17; 71 +/- 4 yr) groups. The inactive groups completed 12 wk (3 d.wk) of aerobic and resistance exercises, and the physically active control groups continued their normal exercise programs. Blood samples were collected before and after the 12-wk period, and the concentrations of serum CRP, plasma <em>interleukin</em>-6 (IL-6), tumor necrosis factor-alpha (TNF-alpha), and <em>interleukin</em>-1 beta (IL-1beta) were determined using separate ELISA.

RESULTS

Control (YPA and OPA) estimated VO2max was unchanged. Exercise training increased estimated VO2max an average of 10.4% and increased strength by an average of 38.1% in both PI groups. Serum CRP decreased with training (YPI and OPI) groups and was not different from the YPA and OPA groups after training. Plasma IL-6 and IL-1beta did not change, whereas TNF-alpha was higher than YPI and YPA at baseline and after the intervention period.

CONCLUSIONS

These results support the use of combined aerobic/resistance training as a modality to reduce the risk of cardiovascular disease development as defined by a decrease in serum CRP concentration in healthy humans.

In the present study, it is demonstrated that cloned surface IgM-positive human B cells can be induced to proliferate and to switch with high frequencies to IgG4 and IgE production after a contact-mediated signal provided by T cell clones and <em>interleukin</em> 4 (IL-4). This T cell signal is antigen nonspecific and is provided by activated CD4+ cells, whereas activated CD8+ or resting CD4+ T cell clones are ineffective. 15-<em>35</em>% of the B cell clones cultured with cloned CD4+ T cells and IL-4 produced antibodies; <em>35</em>-45% of those wells in which antibodies were produced contained IgE and IgG4. In addition to B cell clones that produced IgG4 or IgE only, B cell clones producing multiple isotypes were observed. Simultaneous production of IgG4 and IgE, IgM, IgE, and IgM, or IgG4 and IgE was detected, suggesting that during clonal expansion switching might occur in successive steps from IgM to IgG4 and IgE. In addition, production of only IgM, IgG4, and IgE during clonal expansion indicates that this isotype switching is directed by the way a B cell is stimulated and that it is not a stochastic process.

Conditioned media obtained from fibroblasts cultured from rheumatoid and certain other inflammatory synovia were observed to stimulate [3H]thymidine incorporation in an indicator murine fibroblast line. Synovial fibroblasts derived from the joints of patients with osteoarthritis did not display this property. This effect persisted in culture for many weeks and occurred in the absence of co-stimulatory immune cells. Antibody neutralization studies implicated a role for basic fibroblast growth factor (bFGF), transforming growth factor beta (TGF-beta), granulocyte/macrophage colony-stimulating factor (GM-CSF), and <em>interleukin</em> 1 beta (IL-1 beta) in the increased proliferative activity of synovial fibroblast-conditioned media. Synovial cell synthesis of bFGF, TGF beta 1, GM-CSF, IL-1 beta, and IL-6 was confirmed by <em>35S</em>-methionine labeling and immunoprecipitation. The constitutive production of inflammatory and mitogenic cytokines by synovial fibroblasts may represent the result of long-term, phenotypic changes that occurred in vivo. Persistent cytokine production by synovial fibroblasts may play an important role in the continued recruitment and activation of inflammatory cells in chronic arthritis and in the formation of rheumatoid pannus.

<em>Interleukin</em> 12 (termed IL-12p70 and commonly designated IL-12) is an important immunoregulatory cytokine that is produced mainly by antigen-presenting cells. The expression of IL-12 during infection regulates innate responses and determines the type of adaptive immune responses. IL-12 induces interferon-gamma (IFN-gamma) production and triggers CD4(+) T cells to differentiate into type 1 T helper (Th1) cells. Studies have suggested that IL-12 could play a vital role in treating many diseases, such as viral and bacterial infections and cancers. The unique heterodimeric structure, which IL-12 shares with its family members including IL-23, IL-27, and IL-<em>35</em>, has recently brought more attention to understanding the mechanisms that regulate the functions of IL-12. This article describes the structure and biological activities of IL-12 in both the innate and adaptive arms of the immune system, and discusses the applications of IL-12 in treating and preventing infections.

The pathogenesis of frailty and the role of inflammation is poorly understood. We examined the evidence considering the relationship between inflammation and frailty through a systematic review and meta-analysis. A systematic literature search of papers providing data on inflammatory biomarkers and frailty was carried out in major electronic databases from inception until May 2016. From 1856 initial hits, <em>35</em> studies (32 cross-sectional studies n=3232 frail, n=11,483 pre-frail and n=8522 robust, and 563 pre-frail+robust; 3 longitudinal studies n=3402 participants without frailty at baseline) were meta-analyzed. Cross-sectional studies reported that compared to 6757 robust participants, both 1698 frail (SMD=1.00, 95%CI: 0.40-1.61) and 8568 pre-frail (SMD=0.33, 95%CI: 0.04-0.62) participants had significantly higher levels of C-reactive protein (CRP). Frailty (n=1057; SMD=1.12, 95%CI: 0.27-2.13) and pre-frailty (n=4467; SMD=0.56, 95%CI: 0.00-1.11) were associated with higher serum levels of <em>interleukin</em>-6 compared to people who were robust (n=2392). Frailty and pre-frailty were also significantly associated with elevated white blood cell and fibrinogen levels. In three longitudinal studies, higher serum CRP (OR=1.06, 95%CI: 0.78-1.44,) and IL-6 (OR=1.19, 95%CI: 0.87-1.62) were not associated with frailty. In conclusion, frailty and pre-frailty are associated with higher inflammatory parameters and in particular CRP and IL-6. Further longitudinal studies are needed.

While the role of cytokines in mediating injury during hind limb skeletal muscle ischemia followed by reperfusion has recently been described, the role of cytokines in myocardial infarction and ischemia/reperfusion have remained relatively unexplored. We hypothesize that cytokines play an important role in the regulation of postischemic myocardial inflammation. This study reports the temporal sequence of proinflammatory cytokine gene expression in postischemic/reperfused myocardium and localizes <em>interleukin</em>-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha)-protein by immunostaining. Rats were subjected to either permanent left anterior descending (LAD) occlusion or to <em>35</em> minutes of LAD occlusion followed by reperfusion and sacrificed up to 7 days later. Rat-specific oligonucleotide probes were used to semiquantitatively assess the relative expression of mRNA for TNF-alpha, IL-1 beta, IL-2, IL-6, interferon-gamma (IFN-gamma), and transforming growth factor-beta 1 (TGF-beta 1) utilizing the reverse transcriptase-polymerase chain reaction amplification technique. Increased cardiac mRNA levels for all cytokines except IL-6 and IFN-gamma were measurable within 15 to 30 minutes of LAD occlusion and increased levels were generally sustained for 3 hours. During early reperfusion, mRNA levels for IL-6 and TGF-beta 1 were significantly reduced compared with permanent LAD occlusion. In both groups, cytokine mRNA levels all returned to baseline levels at 24 hours, while IL-1 beta, TNF-alpha, and TGF-beta 1 mRNA levels again rose significantly at 7 days only in animals with permanent LAD occlusion. Immunostaining for IL-1 beta and TNF-alpha protein revealed two patterns of reactivity: 1) microvascular staining for both IL-1 beta and TNF-alpha protein only in postischemic reperfused myocardium in early post-reperfusion time points; and 2) staining of infiltrating macrophages in healing infarct zones which was most prominent at 7 days after permanent LAD occlusion. These results provide evidence for local expression of cytokine mRNA in postischemic myocardium and suggest that regulation of local cytokine release is altered during the postischemic period.

BACKGROUND

Products using the antimicrobial properties of silver nanoparticles (Ag-nps) may be found in health and consumer products that routinely contact skin.

OBJECTIVE

This study was designed to assess the potential cytotoxicity of Ag-nps in human epidermal keratinocytes (HEKs) and their inflammatory and penetrating potential into porcine skin in vivo.

METHODS

We used eight different Ag-nps in this study [unwashed/uncoated (20, 50, and 80 nm particle diameter), washed/uncoated (20, 50, and 80 nm), and carbon-coated (25 and <em>35</em> nm)]. Skin was dosed topically for 14 consecutive days. HEK viability was assessed by MTT, alamarBlue (aB), and CellTiter 96 AQueous One (96AQ). Release of the proinflammatory mediators <em>interleukin</em> (IL)-1beta, IL-6, IL-8, IL-10, and tumor necrosis factor-alpha (TNF-alpha) were measured.

RESULTS

The effect of the unwashed Ag-nps on HEK viability after a 24-hr exposure indicated a significant dose-dependent decrease (p < 0.05) at 0.34 microg/mL with aB and 96AQ and at 1.7 microg/mL with MTT. However, both the washed Ag-nps and carbon-coated Ag-nps showed no significant decrease in viability at any concentration assessed by any of the three assays. For each of the unwashed Ag-nps, we noted a significant increase (p < 0.05) in IL-1beta, IL-6, IL-8, and TNF-alpha concentrations. We observed localization of all Ag-nps in cytoplasmic vacuoles of HEKs. Macroscopic observations showed no gross irritation in porcine skin, whereas microscopic and ultrastructural observations showed areas of focal inflammation and localization of Ag-nps on the surface and in the upper stratum corneum layers of the skin.

CONCLUSIONS

This study provides a better understanding Ag-nps safety in vitro as well as in vivo and a basis for occupational and risk assessment. Ag-nps are nontoxic when dosed in washed Ag-nps solutions or carbon coated.

A preestablished infection with the parasitic helminth, Schistosoma mansoni, significantly reduced the incidence and delayed the onset of experimental autoimmune encephalomyelitis (EAE) in C57BL/6J mice immunized with myelin oligodendrocyte glycoprotein (MOG)(<em>35</em>-55) peptide. The altered disease progression was not solely due to the induction of a strong Th2 response, since intraperitoneal injection of schistosome eggs did not affect disease development. MOG-specific gamma interferon (IFN-gamma), nitric oxide, and tumor necrosis factor alpha production by splenocytes was significantly reduced in schistosome-infected mice compared to uninfected mice. However, similar levels of <em>interleukin</em>-10 (IL-10) were produced in an antigen-specific manner, suggesting that the induction of antigen-specific responses was not inhibited. Analysis of in vivo cytokine production by real-time PCR indicated that IL-12p40, but not IFN-gamma, transcript levels were dramatically reduced in the spinal cords of schistosome-infected, MOG-immunized mice. Furthermore, analysis of the cellular composition of the spinal cords and brains revealed that a preestablished infection with S. mansoni decreased central nervous system (CNS) inflammation, particularly of macrophages and CD4 T cells. These results suggest that schistosomiasis may negatively regulate the onset of EAE by downregulating the production of proinflammatory cytokines and altering CNS inflammation.