Effect of 1,25(OH)2 D3 and 20(OH)D3 on interleukin-1ß-stimulated interleukin-6 and -8 production by human gingival fibroblasts.

BACKGROUND AND OBJECTIVE: Vitamin D-1,25(OH)2 D3 or 1,25D3-maintains healthy osseous tissue, stimulates the production of the antimicrobial peptide cathelicidin and has anti-inflammatory effects, but it can cause hypercalcemia. Evidence links diminished serum levels of 1,25D3 with increased gingival inflammation. Periodontitis progression is associated with increased local production of inflammatory mediators by immune cells and gingival fibroblasts. These include interleukin (IL)-6, a regulator of osteoclastic bone resorption, and the neutrophil chemoattractant IL-8, both regulated by signaling pathways, including NF-κB and MAPK/AP-1. The objectives were to determine the effects of 1,25D3 or a non-calcemic analog, 20-hydroxyvitamin D3 -20(OH)D3 or 20D3-on IL-1ß-stimulated IL-6 and IL-8 production, and NF-κB and MAPK/AP-1 activation, by human gingival fibroblasts. MATERIAL AND METHODS: Human gingival fibroblasts were incubated ± IL-1ß, with or without exposure to 1,25D3 or 20D3. IL-6 and IL-8 in culture supernatants were measured by enzyme-linked immunosorbent assay. NF-κB (p65) and AP-1 (phospho-cJun) and were measured in nuclear extracts via binding to specific oligonucleotides. Data were analyzed using ANOVA and Scheffe's F procedure for post hoc comparisons. RESULTS: IL-1ß-stimulated IL-6 and IL-8 levels were both significantly inhibited (40%-60%) (P<.045) by 1,25D3, but not 20D3 (0%-15% inhibition, not statistically significant). Both 1,25D3 and 20D3 significantly and similarly inhibited IL-1ß-stimulated nuclear levels of p65 and phospho-cJun (P<.02). CONCLUSION: Reduction of the activation of NF-κB and AP-1 alone is not able to inhibit strongly the IL-1ß stimulated IL-6 and IL-8 gene expression. 1,25D3 but not 20D3 may affect some of the many other factors/processes/pathways that in turn regulate the expression of these genes. However, the results suggest that topical application of ligands of the vitamin D receptor may be useful in the local treatment of periodontitis while reducing adverse systemic effects.

Re-evaluating the role of vitamin D in the periodontium.

The importance of vitamin D in maintaining skeletal health via the regulation of calcium has long been recognized as a critical function of this secosteroid. An abundance of literature shows an association between oral bone mineral density and some measure of systemic osteoporosis and suggests that osteoporosis/low bone mass may be a risk factor for periodontal disease. Recently, nonskeletal functions of vitamin D have gained notoriety for several reasons. Many cells that are not associated with calcium homeostasis have been demonstrated to possess membrane receptors for vitamin D. These include activated T and B lymphocytes, and skin, placenta, pancreas, prostate and colon cancer cells. In addition, vitamin D "insufficiency" is a worldwide epidemic and epidemiologic evidence has linked this condition to multiple chronic health problems, including cardiovascular and autoimmune diseases, hypertension and a variety of cancers. Interestingly, there is mounting evidence connecting diminished serum levels of vitamin D with increased gingival inflammation and supporting the concept of "continual vitamin D sufficiency" in maintaining periodontal health. The ability of vitamin D to regulate both the innate and the adaptive components of the host response may play an important role in this process. This review will examine the skeletal and nonskeletal functions of vitamin D, and explore its potential role in protecting the periodontium as well as in regulating periodontal wound healing.

Statins regulate interleukin-1ß-induced RANKL and osteoprotegerin production by human gingival fibroblasts.

BACKGROUND AND OBJECTIVE: Three-hydroxy-3-methyl-glutaryl-CoA (HMG-CoA) reductase competitive inhibitors, or 'statins', are widely used for lowering cholesterol and thereby reducing the risk of a heart attack. Recent data suggest that statins influence metabolic bone activity by their actions on three molecules: RANKL; RANK; and osteoprotegerin (OPG), the soluble decoy receptor for RANKL. The purpose of this study was to evaluate OPG and RANKL production in resting and interleukin-1ß (IL-1ß)-activated human gingival fibroblasts (HGFs), and to determine the effect of statins on their production. MATERIAL AND METHODS: Fibroblasts were pre-incubated with atorvastatin or simvastatin for 24h in serum-free medium, and then incubated with IL-1ß for 6d. The concentration of OPG or RANKL in culture supernatants was measured by specific ELISA. Data were analyzed using analysis of variance and Scheffe's F procedure for post hoc comparison. RESULTS: IL-1ß (1×10(-8) m) stimulated a significant increase in the production of OPG on days 1, 3 and 6. There was a trend towards an increase in RANKL production as a result of stimulation with IL-1ß. Both statins, at multiple concentrations, significantly increased the constitutive RANKL/OPG ratio. Only atorvastatin at the highest concentration (5×10(-6) m) significantly increased the IL-1ß-stimulated RANKL/OPG ratio. CONCLUSION: IL-1ß significantly increased OPG production by HGFs. The statins differed minimally in their effects on OPG and RANKL production by resting and IL-1ß-activated HGFs. Both statins increased constitutive RANKL/OPG ratios, but generally not IL-1ß-stimulated ratios. Thus, statins may influence the production of RANKL and OPG by HGFs to favor bone catabolism, under noninflammatory conditions.

A single-center experience of preemptive anticoagulation for patients with risk factors for allograft thrombosis in renal transplantation.

AIMS: to review our single-center experience of preemptive anticoagulation for the prevention of allograft thrombosis in patients with hypercoagulable states. MATERIAL AND METHODS: this is a retrospective cohort study. Included subjects were first-time kidney allograft recipients transplanted between 2003 and 2007 at a single center, with hypercoagulable states: prior venous thromboembolism, multiple vascular access thromboses, or identifiable thrombophilia. The predictor variable was preemptive anticoagulation and outcome variable was allograft thrombosis. Other risk factors for allograft thrombosis, characteristics of transplantation, and hemorrhagic complications were also examined. RESULTS: among this high-risk cohort (n = 48), 16 received preemptive anticoagulation and 32 did not. The anticoagulated group included significantly more subjects with identifiable thrombophilia (50.0% vs. 0%; p < 0.001). One subject (6.3%) in the anticoagulated group and 6 (18.8%) without anticoagulation developed allograft thrombosis (p = 0.40). A perinephric hematoma was observed in 5 (31.3%) and 2 (6.3%) with and without anticoagulation, respectively (p = 0.03). CONCLUSIONS: preemptive anticoagulation was associated with a non-significant trend towards decreased allograft thrombosis. It may be associated with increased risk of hemorrhage and should be considered cautiously in high-risk patients.

Single-agent lapatinib for HER2-overexpressing advanced or metastatic breast cancer that progressed on first- or second-line trastuzumab-containing regimens.

BACKGROUND: This phase II study evaluated the efficacy and safety of lapatinib in patients with human epidermal growth factor receptor 2 (HER2)-positive advanced or metastatic breast cancer that progressed during prior trastuzumab therapy. PATIENTS AND METHODS: Women with stage IIIB/IV HER2-overexpressing breast cancer were treated with single-agent lapatinib 1250 or 1500 mg once daily after protocol amendment. Tumor response according to RECIST was assessed every 8 weeks. HER2 expression was assessed in tumor tissue by immunohistochemistry and FISH. RESULTS: Seventy-eight patients were enrolled in the study. Investigator and independent review response rates [complete response (CR) or partial response (PR)] were 7.7% and 5.1%, and clinical benefit rates (CR, PR, or stable disease for >or=24 weeks) were 14.1% and 9.0%, respectively. Median time to progression was 15.3 weeks by independent review, and median overall survival was 79 weeks. The most common treatment-related adverse events were rash (47%), diarrhea (46%), nausea (31%), and fatigue (18%). CONCLUSIONS: Single-agent lapatinib has clinical activity with manageable toxic effects in HER2-overexpressing breast cancer that progressed on trastuzumab-containing therapy. Studies of lapatinib-based combination regimens with chemotherapy and other targeted therapies in metastatic and earlier stages of breast cancer are warranted.

Regulation of cytokine production in human gingival fibroblasts following treatment with nicotine and lipopolysaccharide.

BACKGROUND: Patients who smoke are at increased risk for chronic periodontitis (CP). Most studies suggest that the microbial flora in these patients is similar to that found in non-smoking CP patients. Thus, the increased risk for development of CP is not dependent on an altered microbial profile, but rather to some change in the host response to these periopathogens. There is evidence that human gingival fibroblasts (HGF) derived from diseased sites produce greater amounts of interleukin (IL)-6 and IL-8 in vitro than cells derived from healthy sites. This suggests that HGF subpopulations may be selected based upon the inflammatory milieu in which they reside. The hypothesis to be tested was that the combination of nicotine and lipopolysaccharide (LPS) could regulate HGF inflammatory mediator production. METHODS: HGF cell cultures were established from explants derived from 10 patients with CP. HGF cell cultures were stimulated with 1 mM, 1 microM, or 1 nM nicotine +/- Escherichia coli or Porphyromonas gingivalis LPS. At 12, 24, or 48-hour time points, the cells were counted and the supernatant was collected for subsequent IL-6 and IL-8 determination in an enzyme-linked immunosorbent assay. RESULTS: At the 24-hour time point, 1 nM nicotine stimulated IL-6 production compared to control (P=0.02). E. coli LPS alone caused a 3- to 4-fold increase in IL-6 and IL-8 production, whereas P gingivalis LPS did not augment IL-6 or IL-8. A synergistic effect upregulating IL-6 was observed with combined treatment of 1 mM nicotine and E. coli LPS or P gingivalis LPS at the 24-hour time point (P<0.0005 and P=0.002, respectively). Similar effects were seen when IL-8 production was evaluated following HGF stimulation with high doses of nicotine and E. coli LPS or P gingivalis LPS. CONCLUSIONS: These results demonstrate that nicotine by itself can stimulate HGF IL-6 and IL-8 production. Moreover, the combination of high doses of nicotine and either E. coli or P gingivalis LPS can synergistically upregulate cytokine production. These findings support the hypothesis that a proinflammatory fibroblast phenotype may be elicited in an environment enriched with bacterial LPS and nicotine.

Effects of chronic adult periodontitis and endotoxin (LPS) on gingival fibroblast plasma membrane Ca++-pump.

Gingival fibroblasts from patients with chronic adult periodontitis are known to produce cytokines in response to changing levels of bacterial lipopolysaccharide (LPS). Cytokine production is one of numerous cell processes that involve calcium dependent enzymes. It is possible that inflammation may induce changes in the amount of the Ca++-pump protein in gingival fibroblasts which could alter Ca++-dependent activities in these cells including the production and release of cytokines. The purpose of this study was to determine if differences exist in the amount of Ca++-pump protein in the gingival fibroblasts of periodontitis patients relative to control individuals without periodontal disease. Fibroblast explants from healthy tissue and from inflamed tissue from patients with chronic adult periodontitis, grown in culture, were analyzed for quantitative differences in the amount of Ca++-pump protein. Fibroblasts from chronic adult periodontitis patients exhibited significantly lower levels of Ca++-pump protein than fibroblasts from healthy subjects (p=0.0015). However, fibroblasts from chronic adult periodontitis patients, when activated with LPS, did not exhibit significant differences in the amounts of Ca++-pump protein as compared to untreated controls (p = 0.2177). Similarly, cells from healthy subjects did not show significant reduction in Ca++-pump protein following activation with LPS (p = 0.1732). Our results suggest that plasma membrane Ca++-pump is significantly reduced in fibroblasts derived from patients with chronic periodontitis. However, factors other than LPS may be involved in the down-regulation of Ca++-pump protein.

A relationship between proteinase activity and clinical parameters in the treatment of periodontal disease.

The objective of this research was to determine the effectiveness of a biochemical assay which measures proteolytic enzyme activity in gingival crevicular fluid (GCF) and to relate this enzyme activity to clinical parameters traditionally utilized for periodontitis detection. A clinical trial was conducted on 8 periodontitis subjects with > or =4 sites exhibiting a loss of attachment of > or =5 mm and probing depths of > or =5 mm with bleeding on probing. On each subject, a plaque index was performed, followed by GCF sampling at those sites which exhibited a loss of attachment and probing depths. GCF was analyzed for activity against benzoyl-L-arginine-p-nitroanilide in the presence (BAPNA w/gly-gly) and the absence (BAPNA w/o gly-gly) of glycyl-glycine and against MeOSuc-Ala-Ala-Pro-Val-pNA and Suc-Ala-Ala-Pro-Phe-pNA for neutrophil serine proteinases activity (elastase and cathepsin G, respectively). Subsequently, a gingival index was performed, attachment levels and probing depths were recorded using a constant force probe with bleeding on probing being noted. A split-mouth design was employed and half mouths were randomly assigned to the following treatment groups: group A, half of the mouth received scaling/root planing and polishing: group B, half of the mouth received no treatment (control). Subjects were treated, then instructed on toothbrushing and interdental cleaning. After 4 weeks, subjects returned to receive a plaque index; GCF sampling, gingival index, attachment levels, probing depths and bleeding on probing as described above. Using a paired Student t-test, the findings suggest that BAPNA w/gly-gly was significantly less in treatment sites than in non-treated control sites (p=0.05). No such correlation was found for other activities, including neutrophil serine proteinases which were shown to occur in GCF in free, proteolytically active forms. In addition, significant treatment effects were detected for probing depths (p= 0.03) which reduced by 1.3 mm and attachment levels (p=0.02) which gained 0.7 mm. The reduction of P. gingivalis from treated periodontitis sites as detected by a significant decrease in BAPNA w/ gly-gly may prove to be a valuable marker for periodontal disease activity.

HLA-DR alleles are associated with IDDM, but not with impaired neutrophil chemotaxis in IDDM.

Insulin-dependent diabetes mellitus (IDDM) is a risk factor for periodontitis. Depressed neutrophil chemotaxis has been demonstrated in IDDM and in early-onset periodontitis (EOP). HLA-DR antigens are associated with both IDDM and periodontitis. This investigation sought to determine an association of HLA-DR3, -DR4, and -DR53 with impaired neutrophil chemotaxis in an IDDM sample. The neutrophil chemotaxis index of 41 diabetics and 27 controls was determined by a modified Boyden chamber method, and certain class II HLA genotypes were determined by polymerase chain-reaction amplification of genomic DNA by means of sequence-specific primers (PCR-SSP). The mean chemotaxis index of the diabetics was significantly less than that of the controls (p < or = 0.02). HLA-DR3 (p < or = 0.002), -DR4 (p < 0.003), and -DR53 (p < or = 0.001) were associated with IDDM. Neutrophil chemotaxis and glucose metabolism were not significantly correlated. None of the HLA-DR alleles was associated with impaired neutrophil chemotaxis. Therefore, the neutrophil chemotaxis defect of IDDM appears to be independent of these HLA-DR-associated genes.

Interleukin-10 promotes anti-collagen antibody production in type I diabetic peripheral B lymphocytes.

Previous studies have shown that type I diabetes (IDDM) increases the risk of developing periodontitis by 2-3-fold. IDDM patients exhibit destruction of the pancreatic beta cells, most probably caused by an autoimmune reaction. Evidence is accumulating to support the role of the autoimmune response in periodontal pathogenesis. A cytokine, interleukin (IL)-10, has been reported to selectively promote the expansion of a B lymphocyte lineage (CD5/LY1/B1) which has the propensity for secreting high levels of autoantibody. Therefore, the purpose of this project was to evaluate IL-10 production, percentage of CD5 B cells and the frequency of anti-collagen secreting cells in peripheral blood mononuclear cells of age, gender and race matched IDDM patients and controls. IL-10 production was evaluated by an ELISA using the supernatant of adherent peripheral blood cells cultured for 24 h in the presence of Porphyromonas gingivalis lipopolysaccharide (LPS). In 8 of 31 patients, IL-10 levels were significantly increased in IDDM compared to controls and a higher percentage of CD5 B cells was also observed by flow cytometry. In addition, these patients exhibited a higher frequency of anti-collagen secreting cells as elucidated by an ELISPOT. Moreover, treatment with a neutralizing anti-IL-10 antibody diminished the anti-collagen antibody response by 70%. These findings support the concept that a subset of IDDM patients possess an extremely robust IL-10 response following exposure to Gram-negative LPS, which could predispose them to the development of periodontitis through a heightened autoimmune mechanism.

Prostaglandin E2 potentiates interleukin-1 beta induced interleukin-6 production by human gingival fibroblasts.

Increased levels of cytokines and prostanoids have been detected in inflamed gingival tissue and may play an important role in periodontal pathogenesis. Recent studies suggest that monocytic products, such as interleukin (IL)-1 beta, could stimulate IL-6 production by human gingival fibroblasts (HGF). In this context, the production of local cytokines and inflammatory mediators could regulate the secretory capacity of resident gingival fibroblasts. Therefore, the purpose of this study was to determine if PGE2 induced by IL-1 beta could potentiate the IL-6 response by HGF. Utilizing an ELISA, it was determined that maximal IL-6 occurred when HGF were stimulated with 0.10-10 nM IL-1 beta. These concentrations of IL-1 beta also induced a small, but significant increase in PGE2 production by HGF. Interestingly, the combination of IL gamma beta and PGE2 induced a synergistic rise in IL-6 production by HGF. Moreover, inclusion of indomethacin caused a 20% reduction in IL-6 production and totally eliminated PGE2 production. These findings provide additional rationale for the clinical use of NSAIDs in the management of periodontal disease due to their ability to attenuate production of both PGE2, and IL-6. These results suggest the endogenous PGE2 induced by IL-1 beta plays an important regulatory role in IL 6 production by HGF. Moreover, they support the concept that elevated PGE2induced during inflammation can regulate HGF secretory function.

Prostaglandin E2 regulates gingival mononuclear cell immunoglobulin production.

Histological studies have revealed elevated levels of T and B lymphocytes in inflamed gingival tissue. Functional analysis of these B cells has determined that they are spontaneously secreting large amounts of immunoglobulin. Several components of bacterial plaque which accumulate during the onset of periodontal disease induce polyclonal B cell activation, and are most likely responsible for the "hyperactive" state of these gingival B lymphocytes. In addition to this exaggerated humoral response, increased levels of inflammatory mediators, such as prostaglandin (PG) E2, have been implicated in the pathogenesis of disease. Therefore, the purpose of this study was to determine if PGE2 could regulate immunoglobulin production within inflamed gingival tissue. Specimens were harvested during routine surgery of patients with chronic adult periodontitis. Utilizing an ELISA, elevated levels of IgG were detected in the supernatant of cultured gingival mononuclear cells. Inclusion of indomethacin, which inhibits arachidonic acid metabolites such as PGE2, caused a decrease in IgG levels. PGE2 exerted a biphasic effect upon IgG production, with high doses diminishing and low doses increasing IgG levels. From a clinical perspective, these results suggest that elevated levels of PGE2 associated with inflammation will attenuate an IgG response and, as PGE2 production wanes, the local humoral response will rebound. Interestingly, the combination of low dose PGE2 and IL-4 induced a synergistic rise in IgG production. These findings support the theory that local PGE2 levels can regulate immunoglobulin production and potentiate cytokine induced class switching within gingival tissue.

Comparison between mechanical cleaning and an antimicrobial rinse for the treatment and prevention of interdental gingivitis.

This study compared the efficacy of an antimicrobial mouthrinse (0.12% chlorhexidine gluconate) plus toothbrushing (mouthrinse group), mechanical interdental cleaning plus toothbrushing (mechanical group), and toothbrushing alone (control group), at reducing and preventing interdental gingival inflammation. 92 male subjects were examined for interdental inflammation using the Eastman interdental bleeding index at baseline, then monthly for 3 months after using one of the above oral hygiene regimens. The mechanical cleaning group had significant reductions in bleeding sites compared to baseline at 1 month (56.90% versus 13.17%) that persisted throughout the study (2 months = 6.65%, 3 months = 5.70%). The other regimens showed no significant bleeding reduction at any time point in the study. The mechanical interdental cleaning group showed improvement over baseline at 1 month with the full benefit apparent after 2 months. The effect of location in the mouth on bleeding reduction was also assessed. The % of posterior sites which bled was always higher than anterior sites. Analysis of maxillary versus mandibular, and buccal versus lingual sites showed no significant differences. Additional observations of the data demonstrated that sites which bled at baseline were more likely to stop bleeding in the mechanical cleaning group. Also, sites which did not bleed at baseline were unlikely to bleed subsequently when mechanical cleaning was used. Neither of these observations were true for the other cleaning regimens. These data show that only mechanical interdental plaque removal combined with toothbrushing is effective at reducing or preventing interdental inflammation. This underscores the importance of instituting mechanical interdental cleaning to eliminate interdental inflammation.

Anti-class II antibodies potentiate IgG2a production by lipopolysaccharide-stimulated B lymphocytes treated with prostaglandin E2 and IFN-gamma.

IFN-gamma secretion by Th1 cells has been shown to preferentially promote the production of IgG2a in LPS-stimulated murine B lymphocytes. We recently reported that PGE2 potentiated the ability of IFN-gamma to augment IgG2a production in both Ag-specific and polyclonal systems via a cAMP-dependent pathway. Because antibodies (Ab) directed against class II MHC molecules have been shown to induce a rise in B cell cAMP, we hypothesized that this event, like PGE2 treatment, would promote the production of IgG2a. In this manuscript, cultures of small and large B cells treated with anti-Ia Ab are shown to produce significantly higher levels of IgG2a, compared with cultures treated only with IFN-gamma and LPS. Moreover, the combined treatment of B lymphocytes with IFN-gamma and PGE2 followed by anti-Ia and LPS resulted in a fourfold rise in IgG2a levels compared with IFN-gamma and LPS. Only anti-class II, but not anti-class I Ab, stimulated IgG2a production. Utilizing an ELISA spot assay, the frequency of IgG2a-secreting B cells was determined to be elevated fourfold in anti-Ia treated B cells. B cell cultures incubated with either PGE2 or anti-Ia exhibited elevated levels of cAMP and treatment with IFN-gamma primed these lymphocytes to the cAMP-elevating effects of either PGE2 or anti-Ia. Finally, RpcAMP, a cAMP antagonist that blocks cAMP from activating protein kinase A, prevented the increased production of IgG2a induced by anti-Ia Ab. These results support the theory that a cAMP pathway exists that promotes B cell IgG2a production. Within this pathway, IFN-gamma sensitizes B lymphocytes to cAMP elevators such as anti-class II Ab, and in conjunction with LPS, causes an increase in the frequency of IgG2a-secreting cells and the amount of IgG2a produced. These observations suggest that, after exposure to viral Ag in vivo, interaction between IFN-gamma-primed murine B cells and T cells will potentiate production of IgG2a, the predominant murine anti-viral Ig.

Examiner reliability for an invasive gingival bleeding index.

This investigation was undertaken to determine the intra- and inter-examiner reliability of the method of stimulation for bleeding used in the Eastman interdental bleeding index. 26 subjects were examined twice, 1 h apart, by either a single examiner or 2 examiners in each half of their mouths, for the presence bleeding after stimulation with a wooden interdental cleaner. Scores were tabulated and intra- and inter-examiner % agreements and kappa-coefficients calculated. Z-tests were performed on the pairs of agreement statistics to check for significant differences. Overall, intra-examiner agreement statistics were high (91.3% to 93.1% agreement; 0.79 to 0.86 kappa-coefficient). Further breakdowns of the data into facial and lingual sites by arch and location (anterior or posterior) resulted in similar levels of reliability, with no significant differences within examiners. The overall inter-examiner agreement statistics were good (82.8% to 87.6% agreement; 0.62-0.75 kappa coefficient). When inter-examiner data were analyzed at facial or lingual sites by arch and location, a significant difference existed in reliability for mandibular posterior lingual sites, but reliability was high in all other areas. These data demonstrate a high level of reproducibility for this method, which suggests that the Eastman interdental bleeding index is suitable for clinical trials and epidemiologic studies of interdental gingivitis.

A new view of prostaglandin E regulation of the immune response.

Prostaglandins, particularly those of the E series, are widely regarded as immunosuppressive products of eukaryotic cells that can downregulate many aspects of B- and T-cell function. In this article, Richard Phipps and colleagues present a different concept of E series prostaglandins, based on recent evidence supporting a role for prostaglandins as potentiators of immunoglobulin class switching and of the synthesis of selected cytokines and cytokine receptors.

Antigen-specific IGG2A production in response to prostaglandin E2, immune complexes, and IFN-gamma.

Immune complexes (IC) can inhibit the differentiation of B lymphocytes into IgM secreting plasma cells in both Ag-specific and polyclonal systems. This report describes the ability of IC and IFN-gamma, or IFN-gamma and PGE2 to regulate the class of Ig produced in an Ag-specific system. In an in vitro model system using fluorescein (FL)-Brucella abortus as Ag, we previously showed that IC composed of an anti-FL antibody and FL-Ag inhibited the ability of FL-specific B cells to develop into IgM plaque-forming cells. In addition, PGE2 sensitized resting B cells to IC, whereas pretreatment with IL-4 alleviated the IC-mediated decrease in the IgM anti-FL response. In this manuscript, we demonstrate that IFN-gamma has antagonistic effects compared to IL-4, and potentiates the IC-induced decrease in the IgM antibody response. Interestingly, we discovered that the virtual ablation of the anti-FL IgM response exhibited by B cells treated with IC and IFN-gamma was accompanied by a dramatic increase in the anti-FL IgG2a response. Furthermore, resting B lymphocytes pulsed with IFN-gamma and PGE2, but not PGF2 alpha, exhibited augmented IgG2a production. This effect was also observed when IFN-gamma-pulsed B cells were stimulated with other agents, such as dibutyryl cAMP and cholera toxin, that elevate intracellular levels of cAMP. In addition, RpcAMP, the R-isomer of a sulfur-modified cAMP and a cAMP antagonist, inhibited anti-FL IgG2a responses. The ability of cAMP elevators such as PGE2 to promote IgG2a production, as well as to increase the frequency of IgG2a secreting cells, was demonstrated in B lymphocytes treated with IFN-gamma and polyclonally activated by LPS. Overall, our results demonstrate that IFN-gamma and PGE2-treated B lymphocytes challenged with antigen or LPS generate elevated levels of IgG2a via a cAMP-dependent pathway. These observations suggest that in vivo, PGE2 secreted by cells such as macrophages potentiates the ability of IFN-gamma to promote an IgG2a response, the predominant murine antiviral Ig.

A dynamic interregional theory of migration and population growth.

PIP: The authors attempt to bring the theory of regional amenity value determination together with a theoretical framework on migration to form a cohesive theoretical model. They build on previous efforts to do this by introducing factors such as the housing market and determination of the equilibrium level of real full income. The implied geographical focus is on the United States.^ieng

Elevated levels of intracellular cAMP sensitize resting B lymphocytes to immune complex-induced unresponsiveness.

The ability of immune complexes (IC) to regulate B lymphocyte differentiation was investigated. Using an in vitro model, we previously demonstrated that macrophages (M phi) or lymphoid dendritic cells pulsed with IC differentially regulated B cell function, inducing unresponsiveness or stimulation, respectively. The capacity of M phi to induce unresponsiveness was dependent upon two signals, an antigen-specific one supplied by the IC and M phi-secreted prostaglandin (PG)E2. Total inhibition of antibody production was never achieved as a small percentage of B lymphocytes were resistant to IC-induced unresponsiveness. In this study, utilizing an accessory cell-free system, we demonstrate that splenic B cell fractions separated on Percoll density gradients are heterogeneous in their sensitivity to IC-mediated unresponsiveness. Small resting B lymphocytes are exquisitely sensitive to IC-mediated negative signaling and exhibit virtual total ablation of antibody responses. Conversely, large activated B cells are more refractory to this inhibitory pathway. PGE2 and other agents which elevate cAMP potentiate IC-induced unresponsiveness in resting, but not activated B lymphocytes. In addition treatment of resting B cells with PGF2 alpha, which did not elevate cAMP, failed to sensitize these cells to IC-mediated negative signaling. Unresponsiveness induced by IC is selective for specific aspects of B lymphocyte activation, since B cell differentiation but not proliferation is affected. Furthermore, pre-treatment of resting B lymphocytes with interleukin 4 prevents the IC-induced ablation of IgM antibody responses. Overall, our results indicate that the binding of IC by resting B lymphocytes provides a potent mechanism for inhibiting differentiation without affecting proliferation. These observations suggest that in vivo, IC play an important role in regulating the memory B lymphocyte pathway.