Production Of IL-1alpha Research Articles

Bone Marrow Microenvironment As a Potential Key Regulator of Hematopoietic Cells Stemness in Myelodysplastic Syndromes (MDS)

Background and Rationale. Nuclear Phospholipase C (PLC) beta1 plays a pivotal role in Myelodysplastic Syndromes (MDS) and Azacytidine response [Cocco, L., et al. J Lipid Res. 2015; 56: 1853-60]. Hematopoietic Stem Cells (HSCs) primarily reside within the bone marrow (BM) microenvironment, where mesenchymal stromal cells (MSCs) regulate hematopoiesis through direct interactions with HSCs or cytokine secretion [Blau O, et al. Blood. 2011; 118:5583-92]. The aim of this study was to better elucidate the role of the BM microenvironment in interacting with MDS/AML cells, with particular emphasis on investigating the correlation between PLCbeta1 modulation and the distinctive BM microenvironment response after co-culture, in terms of PLCs gene expression, myeloid differentiation markers and cytokine secretion. Patients and Methods. As it is a very preliminary study, here we analyzed mononuclear cells obtained from 4 higher-risk MDS (RAEB-1, R-IPSS High or Very High, Complex Karyotype), who gave informed consent in accordance with the Declaration of Helsinki. Two patients were treated with Azacytidine (AZA, 75 mg/mq/die for 7 days every 28 days) and one of them reached the fifth cycle with a stable disease. All samples were obtained from the IRCCS-Institute of Hematology “L. e A. Seràgnoli”, Bologna, Italy, where clinical evaluations were also conducted. Co-cultures were established with HS-5 MSCs, while Real-Time PCR results were normalized using molecular data obtained from 10 healthy donors (HD). PLCbeta1 overexpression/silencing in vitro model was represented by THP-1 monocytic cells subjected to lentiviral transfection. Both wild-type and PLCbeta1 overexpressing/silencing cells (THP-1 OV/KD) were co-cultured with HS-5 cells for 96 hours. Results. Patients showed distinct basal expression patterns of PLCbeta1: higher in 1/4 MDS and lower in 3/4 MDS, as compared to HD. Irrespective of PLCbeta1 basal expression levels, or response to AZA, in 3/4 patients CD11b increased and CD14 decreased after co-culture. Only one patient, characterized by being a therapy-related MDS, displayed a reduced expression in all myeloid differentiation markers during co-culture. During AZA therapy and after co-culture, one patient showing a stable disease had a significant PLCbeta1 reduction, CD11b increase and CD14 decrease. In THP-1 OV cells co-cultured with HS-5 cells, we observed CD11b reduction, while no significant differences in both PLCs and other myeloid differentiation markers. In contrast, THP-1 KD cells, following co-culture, exhibited a notable increase in CD11b and a decreased CD14 expression. Moreover, THP-1 OV cells alone displayed inhibition of IL-8 secretion, that disappeared after co-culture, in favour of IL-1beta secretion. THP-1 KD cells exhibited suppressed secretion of IL-1alpha, IL-1beta, and IL-8. However, during co-culture, the production of both IL-1alpha and IL-1beta was restored. Conclusions. Our results, that must be confirmed in a larger group of patients, show that the impact of co-culture is not associated with the baseline levels of PLCbeta1, but co-culture recapitulates the PLCbeta1 KD in vitro model and the lack of response to AZA, showing an increase of CD11b and a CD14 reduction. Moreover, the presence of MSCs reversed cytokine secretion, as in co-cultured THP1 KD cells IL-1alpha and IL-1beta were highly secreted again. Considering their multiple roles in both physiological and pathological conditions, ongoing analyses are now further investigating their role in BM microenvironment, also using MDS-MSCs. In addition, we are now exploring the involvement of BM microenvironment in maintaining leukemic cells in a more undifferentiated state and the potential implications of these mechanisms on AZA response/resistance.

Altered monocyte phenotype and dysregulated innate cytokine responses among people living with HIV and opioid-use disorder.

Opioid-use disorders (OUD) and hepatitis C or B co-infection (HEP) are common among people living with HIV (PLHIV). The impact of OUD on innate and adaptive immunity among PLHIV with and without HEP is unknown. To investigate the impact of OUD on monocyte and T-cell phenotypes, cytokine responses to lipopolysaccharide (LPS) and phytohemagglutinin (PHA), and plasma inflammatory markers, among PLHIV with and without HEP. Cross-sectional study enrolling PLHIV receiving ART, with and without OUD. Flow cytometry determined monocyte and T-cell phenotypes; LPS and PHA-induced cytokine production was assessed following LPS and PHA stimulation by multiplex cytokine array; plasma IL-6, soluble CD163, and soluble CD14 were measured by ELISA. Twenty-two PLHIV with OUD and 37 PLHIV without OUD were included. PLHIV with OUD exhibited higher frequencies of intermediate (CD14CD16) and nonclassical (CD14CD16) monocytes when compared with PLHIV without OUD (P = 0.0025; P = 0.0001, respectively), regardless of HEP co-infection. Soluble CD163 and monocyte cell surface CD163 expression was increased among PLHIV with OUD and HEP, specifically. Regardless of HEP co-infection, PLHIV with OUD exhibited reduced production of IL-10, IL-8, IL-6, IL-1alpha, and TNF-alpha in response to LPS when compared with PLHIV without OUD; PHA-induced production of IL-10, IL-1alpha, IL-1beta, IL-6, and TNF-alpha were also reduced among individuals with OUD. OUD among PLHIV are associated with altered monocyte phenotypes and a dysregulated innate cytokine response. Defining underlying mechanisms of opioid-associated innate immune dysregulation among PLHIV should be prioritized to identify optimal OUD treatment strategies.

Safety Assessment of Nano-Hydroxyapatite as an Oral Care Ingredient according to the EU Cosmetics Regulation

Hydroxyapatite nanoparticles (HAP-NP) are incorporated in oral care products such as toothpastes and mouthwashes to treat dental sensitivity or to promote enamel remineralisation. Despite the good performance of HAP-NP in this application, it is important to ensure its safety for consumers. For that reason, the Scientific Committee on Consumer Safety (SCCS) evaluated the safety of HAP-NP as an oral care ingredient, but the issued opinion was not completely conclusive and the SCCS recommended that additional tests should be performed. Here, we used a commercially available human gingival epithelium (HGE) as a non-animal alternative and MTT cell viability, LDH activity, and IL-1alpha production were evaluated after 3.1% HAP-NP treatment for 10 min, 1 h, and 3 h. Moreover, the absorption of HAP-NP in the gingival tissue was assessed by transmission electron microscopy (TEM) analysis. Finally, the dissolution behaviour of HAP-NP in simulated gastric fluid was also investigated. No deleterious effect was observed for HGE tissues incubated with HAP-NP for all time-points and parameters evaluated. Moreover, a complete dissolution of 3.1% HAP-NP in simulated gastric fluid was observed after 7.5 min at 37 °C. In conclusion, our results evidence the safety of HAP-NP for oral care products with the use of an in vitro replacement alternative for human gingival epithelium and a simulated gastric fluid assay.

Monocyte Phenotype and Polyfunctionality Are Associated With Elevated Soluble Inflammatory Markers, Cytomegalovirus Infection, and Functional and Cognitive Decline in Elderly Adults.

Monocytes are mediators of the inflammatory response and include three subsets: classical, intermediate, and nonclassical. Little is known about the phenotypical and functional age-related changes in monocytes and their association with soluble inflammatory biomarkers, cytomegalovirus infection, and functional and mental decline. We assayed the activation ex vivo and the responsiveness to TLR2 and TLR4 agonists in vitro in the three subsets and assessed the intracellular production of IL1-alpha (α), IL1-beta (β), IL-6, IL-8, TNF-α, and IL-10 of elderly adults (median 83 [67-90] years old;n= 20) compared with young controls (median 35 [27-40] years old;n= 20). Ex vivo, the elderly adults showed a higher percentage of classical monocytes that expressed intracellular IL1-α (p= .001), IL1-β (p= .001), IL-6 (p= .002), and IL-8 (p= .007). Similar results were obtained both for the intermediate and nonclassical subsets and in vitro. Polyfunctionality was higher in the elderly adults. The functionality ex vivo was strongly associated with soluble inflammatory markers. The activation phenotype was independently associated with the anti-cytomegalovirus IgG levels and with functional and cognitive decline. These data demonstrate that monocytes are key cell candidates for the source of the high soluble inflammatory levels. Our findings suggest that cytomegalovirus infection might be a driving force in the activation of monocytes and is associated with the functional and cognitive decline.

Immune mechanisms in periodontitis � effect of IL-4 gene polymorphisms on cytokine production

Introduction: Periodontitis is a chronic inflammatory disease of the teeth-supporting tissues in which genetic predisposition, dental plaque bacteria and immune mechanisms play important roles. The aim of this study was to assess the influence of IL-4 gene polymorphisms on cytokine production Methods: Cytokine production (IL-1alpha, IL-1beta, IL-4, IL-5, IL-6, IL-8 , IL-10, TNF–alpha, INF–gamma) was studied after in vitro stimulation of isolated peripheral blood mononuclear cells (PBMC) by mitogens and dental plaque bacteria (Aggregatibacter actinomycetemcomitans, Escherichia coli, Tannerrella forsythensis, Porphyromonas gingivalis and Heat Shock Protein 60). Cytokine production was detected by multiplex cytokine analysis system Luminex. The results were correlated with IL-4 genotypes in patients with chronic periodontitis (CP) and healthy controls. Results: We found that mononuclear cells isolated from peripheral blood of CP patients with investigated IL-4 polymorphisms (-589C/T, -33C/T, VNTR in intron 3) significantly changed production of IFN-gamma, IL-10, IL-1alpha and IL-1beta, TNF-alpha, IL-10, MCP-1, IL-6 after stimulation by all selected bacteria and HSP (p<0.05). Conclusion: In patients with CP, IL-4 polymorphisms may influence the function of mononuclear cells to produce not only IL-4 but also other cytokines. The results of this study support Seymour’s hypothesis (1999) about the role of TH2 produced cytokines such as IL-4 in genetic predisposition to chronic periodontitis.

O041 Predominant role of macrophage-derived IL-1alpha but not IL-1beta in atherosclerosis

Introduction Atherosclerosis is a chronic inflammatory condition affecting the arterial vasculature. Its clinical manifestations, such as myocardial infarction and ischemic stroke, are the leading causes of morbidity and mortality worldwide. IL-1 receptor (IL-1R)-signaling severely exacerbates the chronic vascular inflammation that drives disease progression. While the contribution of inflammasome-activation to atherogenesis in vivo remains controversial, IL-1beta is currently targeted in clinical trials. However, as both IL-1alpha and IL-1beta signal via IL-1R and exhibit overlapping functions, the relative contribution of each IL-1 isoform to atherogenesis is still unclear. Methods We have compared the impact of both IL-1 isoforms on disease progression in mouse models of atherosclerosis, and have investigated the mechanisms of IL-1alpha versus IL-1beta induction by different atherogenic lipids in macrophage-foam cells. Results Here we report that deficiency in hematopoietic IL-1alpha markedly reduced atherosclerosis by 51%, whereas no significant effect was observed in mice lacking BM-derived IL-1beta, suggesting that macrophage-derived IL-1alpha - and not IL-1beta - represents the pro-atherogenic IL-1 isoform. Moreover, our study identified the fatty acid oleic acid, which is abundant amongst plaque lipids, as a selective trigger of potent IL-1alpha - but not IL-1beta - responses in foam cells. This atherogenic IL-1alpha production was independent of inflammasome activation, but surprisingly required the cellular expression of pro-IL-1beta. Hence, IL-1beta−/− macrophages were unable to secrete IL-1alpha, which may explain the conflicting reports showing reduced atherosclerosis in IL-1beta-deficient mice. Furthermore, feeding of an oleate-enriched but cholesterol-free diet aggravated atherogenesis, suggesting that IL-1alpha is both necessary and sufficient to induce the development of atherosclerotic plaques. Conclusion Taken together, our data demonstrate that macrophage-foam cells promote atherogenesis primarily through production of IL-1alpha, and therefore suggest that the role of inflammasome activation and IL-1beta in atherosclerosis may need to be reconsidered. Most importantly, they indicate that IL-1alpha and not IL-1beta should be targeted therapeutically in patients with cardiovascular disease.

MTHPC‐mediated Photodynamic Treatment Up‐regulates the Cytokines VEGF and IL‐1alpha

Photodynamic therapy (PDT) of cancer induces oxidative stress, which intervenes in the expression of cytokines by tumor cells. The cytokines might have either a positive or a negative impact on tumor eradication. Here, we studied the expression of cytokines vascular endothelial growth factor (VEGF) and interleukin-1alpha (IL-1alpha) in the human epidermoid carcinoma A-431 cells following m-tetra(3-hydroxyphenyl)-chlorin (mTHPC)-mediated PDT in vitro and assessed the IL-1alpha effect on VEGF expression. Quantitative polymerase chain reaction and enzyme-linked immunosorbent assay revealed the enhanced production of VEGF and IL-1alpha both on mRNA and protein levels by mTHPC-loaded cells after light exposure. The silencing of IL1A by small interfering RNA resulted in decreased production of IL-1alpha and a reduced amount of VEGF. Furthermore, exogenous recombinant IL-1alpha stimulated the VEGF expression after PDT. Thus, in addition to the cytotoxic action on the A-431 cells, mTHPC-mediated PDT stimulated the production of VEGF and IL-1alpha, and IL-1alpha contributed to the VEGF overexpression. These data establish IL-1alpha as a possible target of combined cancer treatment.

Peroxisome-proliferator-activated receptors γ and peroxisome-proliferator-activated receptors β/δ and the regulation of interleukin 1 receptor antagonist expression by pioglitazone in ischaemic brain

The imbalance between the production and release of interleukin-1 (IL-1) ligands, IL-1alpha, IL-1beta and IL-1 receptor antagonist (IL-1ra) in ischaemic brain exaggerates inflammatory responses and contributes to neuronal death. Cerebral ischaemia also upregulates the peroxisome-proliferator-activated receptor (PPAR) gamma. We studied in rats the effects of the PPARgamma agonist, pioglitazone, on the regulation of IL-1beta, IL-1ra and IL-1 receptor I (IL-1RI) expression in ischaemic brain after occlusion of the middle cerebral artery for 90 min. Pioglitazone or vehicle was infused intracerebroventricularly over a 5-day period before, during and 24 or 48 h after middle cerebral artery occlusion. The expression of IL-1beta, IL-1ra and IL-1RI in the peri-infarct cortex was investigated by immunohistochemistry, Western blotting and immunofluorescence staining. The mechanisms of the IL-1ra regulation by pioglitazone and the neuroprotection under excitotoxic neuronal injury were studied in primary cortical neurones expressing PPARgamma and PPAR beta/delta. Cerebral ischaemia increased the expression of IL-1beta, IL-1RI and IL-1ra in the ischaemic cortex. Pioglitazone reduced IL-1beta, but upregulated IL-1ra and increased the number of IL-1ra immunoreactive cells. In primary cortical neurones, pioglitazone stimulated the IL-1ra production via activation of the PPARbeta/delta, but prevented excitotoxic neuronal injury and death by a PPARgamma-dependent mechanism. Our data demonstrate that activation of PPARgamma and PPAR beta/delta by proglitazone in neurones triggers diverse neuroprotective mechanisms. The restoration of the equilibrium between I1-1beta and IL-1ra in ischaemic brain tissue limits IL-1beta signalling, reduces inflammatory responses and is an important mechanism by which thiazolidinediones improve the recovery from ischaemic stroke.

Upregulation of interleukin‐1 by Epstein–Barr virus latent membrane protein 1 and its possible role in nasopharyngeal carcinoma cell growth

Nasopharyngeal carcinoma (NPC) is associated with Epstein-Barr virus (EBV) infection. We previously found that interleukin (IL)-1alpha and IL-1beta significantly increased in NPC tissues. This study investigated what EBV-encoded proteins were involved in such IL-1 production. IL-1alpha and IL-1beta messenger ribonucleic acids (mRNAs) were detected in the EBV latent membrane protein 1 (LMP1) transfectant (LMP135) only by reverse transcriptase-polymerase chain reaction (RT-PCR). LMP1-mediated IL-1alpha and IL-1beta production could be enhanced by tumor necrosis factor alpha (TNF-alpha), determined by enzyme-linked immunosorbent assay (ELISA). Moreover, IL-1alpha and IL-1beta mRNAs and proteins were increased in a dose-dependent manner in epithelial cells transiently transfected by an LMP1 plasmid. Besides, immortalized human epidermal keratinocyte (RHEK-1) epithelial cells could be enhanced to proliferate by IL-1alpha and IL-1beta determined by water-soluble tetrazolium salt (WST-1) assay. EBV LMP1 is capable of upregulating IL-1alpha and IL-1beta secretions from epithelial cells and positively modulated by TNF-alpha. This may consequently contribute to tumor growth in patients with NPC.

Listeriolysin O-Dependent Bacterial Entry into the Cytoplasm Is Required for Calpain Activation and Interleukin-1α Secretion in Macrophages Infected withListeria monocytogenes

Listeriolysin O (LLO), an hly-encoded cytolysin of Listeria monocytogenes, plays an essential role in the entry of L. monocytogenes into the host cell cytoplasm. L. monocytogenes-infected macrophages produce various proinflammatory cytokines, including interleukin-1 alpha (IL-1 alpha), that contribute to the host immune response. In this study, we have examined IL-1 alpha production in macrophages infected with wild-type L. monocytogenes or a nonescaping mutant strain deficient for LLO (Delta hly). Expression of IL-1 alpha mRNA and accumulation of pro-IL-1 alpha in the cytoplasm were induced by both strains. In contrast, the secretion of the mature form of IL-1 alpha from infected macrophages was observed in infection with wild-type L. monocytogenes but not with the Delta hly mutant. A recovery of the ability to induce IL-1 alpha secretion was shown in a mutant strain complemented with the hly gene. The Toll-like receptor (TLR)/MyD88 signaling pathway was exclusively required for the expression of pro-IL-1 alpha, independently of LLO-mediated cytoplasmic entry of L. monocytogenes. The LLO-dependent secretion of mature IL-1 alpha was abolished by addition of calcium chelators, and only LLO-producing L. monocytogenes strains were able to induce elevation of the intracellular calcium level in infected macrophages. A calcium-dependent protease, calpain, was implicated in the maturation and secretion of IL-1 alpha induced by LLO-producing L. monocytogenes strains based on the effect of calpain inhibitor. Functional activation of calpain was detected in macrophages infected with LLO-producing L. monocytogenes strains but not with a mutant strain lacking LLO. These results clearly indicated that LLO-mediated cytoplasmic entry of bacteria could induce the activation of intracellular calcium signaling, which is essential for maturation and secretion of IL-1 alpha in macrophages during L. monocytogenes infection through activation of a calcium-dependent calpain protease. In addition, recombinant LLO, when added to macrophages infected with the Delta hly strain, could induce calcium influx and IL-1 alpha secretion at doses exhibiting cytolytic activity, suggesting that LLO produced by intracellular L. monocytogenes may be implicated in induction of calcium influx through pore formation.

Spontaneous production of interleukin-1 alpha and beta by alveolar cells from patients with sarcoidosis

The spontaneous production of interleukin-1 alpha and beta by alveolar cells obtained by BAL from 7 active pulmonary sarcoidosis and 5 normal volunteers was evaluated. The activity of disease in one case was considered to be highly active because of the chest X-ray pattern (diffuse micronodular shadows), highly intense Ga up take in lungs, increased number of BAL cells and high level of S-ACE. The contents of IL-1 alpha and beta were measured by ELISA in the culture supernatants of alveolar cells after 24 hours culture without any stimulus. Large amounts of IL-1 alpha and beta production were found in highly active case. No significant amount of IL-1 alpha and beta, however, was detected in 6 other active sarcoidosis cases and 5 normal volunteers. Therefore, spontaneous release of IL-1 alpha and beta in vitro from alveolar cells in sarcoidosis might be considered as an index for the necessity of systemic corticosteroid treatment and its relationship to the spontaneous remission of sarcoidosis in Japanese patients was discussed.

Development of an in vitro model for radiation-induced effects on oral keratinocytes

Changes in epithelial cell activity and the production of pro-inflammatory cytokines were examined utilizing an organotypic culture system as an in vitro model to study the effects of radiation on oral keratinocytes to simulate what is thought to occur in radiation-induced oral mucositis. Monolayer cultures of oral keratinocyte were irradiated by varying the dose. Cell injury was assessed using a colony forming efficiency (CFE) assay. Third passage oral keratinocytes were seeded onto AlloDerm to form a 3D construct of an ex vivo produced oral mucosa equivalent (EVPOME) which was irradiated with 0, 1, 3 and 8Gy. Formalin-fixed sections of the EVPOME were used for histology and immunohistochemistry to examine proliferative capacity. Epithelial cell viability of EVPOME was measured by MTT assay. Spent culture medium was used to determine post-radiation pro-inflammatory cytokine production. Basal cells became more swollen and pyknotic as radiation increased, implying loss of cell viability also determined by MTT assay. The number of Ki-67 immunopositive cells and CFE showed negative correlation with radiation, indicating loss of cell proliferative capacity. The production of pro-inflammatory cytokines, IL-1alpha and IL-8, tended to increase in a radiation dose dependent manner. The EVPOME lacking submucosal cellular components was a useful model.

Calcium Channel Blockers Differentially Modulate Cytokine Production by Peripheral Blood Mononuclear Cells

Calcium channel blockers (CCB) are known to modulate immune reactions, so the present study was performed to examine the effects of various CCBs that have shown different effects on transcription factors and on the production of pro-inflammatory cytokines by human peripheral blood mononuclear cells (PBMC). PBMC from healthy volunteers were isolated by Ficoll-paque density centrifugation. To study the effect of CCBs, the PBMC were stimulated with lipopolysaccharide or concanavalin A. After 24 h of incubation, the supernatants were harvested and the interleukin (IL)-1alpha, -1beta, and -6, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma levels were determined by specific enzyme-linked immunosorbent assay. The production of IL-1alpha and -1beta stimulated with lipopolysaccharide was significantly increased in the presence of amlodipine. In contrast, nifedipine and verapamil suppressed the production of IL-1beta, TNF-alpha, and IFN-gamma. Amlodipine and diltiazem significantly increased production of IL-1alpha stimulated with concanavalin A. Nifedipine inhibited production of IL-1alpha, IL-6, and IFN-gamma. Verapamil suppressed production of IFN-gamma. Differential modulation of cytokine production was seen with various CCBs, and the suppressive effect of nifedipine was most prominent.

Mactinin, a fragment of cytoskeletal α-actinin, is a novel inducer of heat shock protein (Hsp)-90 mediated monocyte activation

BackgroundMonocytes, their progeny such as dendritic cells and osteoclasts and products including tumor necrosis factor (TNF)-α, interleukin (IL)-1α and IL-1β play important roles in cancer, inflammation, immune response and atherosclerosis. We previously showed that mactinin, a degradative fragment of the cytoskeletal protein α-actinin, is present at sites of monocytic activation in vivo, has chemotactic activity for monocytes and promotes monocyte/macrophage maturation. We therefore sought to determine the mechanism by which mactinin stimulates monocytes.ResultsRadiolabeled mactinin bound to a heterocomplex on monocytes comprised of at least 3 proteins of molecular weight 88 kD, 79 kD and 68 kD. Affinity purification, mass spectroscopy and Western immunoblotting identified heat shock protein (Hsp)-90 as the 88 kD component of this complex. Hsp90 was responsible for mediating the functional effects of mactinin on monocytes, since Hsp90 inhibitors (geldanamycin and its analogues 17-allylamino-17-demethoxygeldanamycin [17-AAG] and 17-(dimethylaminoethylamino)-17-demethoxygeldanamycin [17-DMAG]) almost completely abrogated the stimulatory activity of mactinin on monocytes (production of the pro-inflammatory cytokines IL-1α, IL-1β and TNF-α, as well as monocyte chemotaxis).ConclusionMactinin is a novel inducer of Hsp90 activity on monocytes and may serve to perpetuate and augment monocytic activation, thereby functioning as a "matrikine." Blockage of this function of mactinin may be useful in diseases where monocyte/macrophage activation and/or Hsp90 activity are detrimental.

Effect of Lactobacillus rhamnosus GR‐1 and Lactobacillus reuteri RC‐14 on the ability of Candida albicans to infect cells and induce inflammation

Vulvovaginal candidiasis, a high prevailing infection worldwide, is mainly caused by Candida albicans. Probiotic Lactobacillus reuteri RC-14 and Lactobacillus rhamnosus GR-1 have been previously shown to be useful as adjuvants in the treatment of women with VVC. In order to demonstrate and better understand the anti-Candida activity of the probiotic microorganisms in an in vitro model simulating vaginal candidiasis, a human vaginal epithelial cell line (VK2/E6E7) was infected with C.albicans 3153a and then challenged with probiotic L. rhamnosus GR-1 and/or L. reuteri RC-14 or their respective CFS (alone or in combination). At each time point (0, 6, 12 and 24 hr), numbers of yeast, lactobacilli and viable VK2/E6E7 cells were determined and, at 0, 6 and 12 hr, the supernatants were measured for cytokine levels. We found that C. albicans induced a significant increase in IL-1alpha and IL-8 production by VK2/E6E7 cells. After lactobacilli challenge, epithelial cells did not alter IL-6, IL-1alpha, RANTES and VEGF levels. However, CFS from the probiotic microorganisms up-regulated IL-8 and IP-10 levels secreted by VK2/E6E7 cells infected with C. albicans. At 24 hr of co-incubation, L. reuteri RC-14 alone and in combination with L. rhamnosus GR-1 decreased the yeast population recoverable from the cells. In conclusion, L. reuteri RC-14 alone and together with L. rhamnosus GR-1 have the potential to inhibit the yeast growth and their CFS may up-regulate IL-8 and IP-10 secretion by VK2/E6E7 cells, which could possibly have played an important role in helping to clear VVC in vivo.

Effects of endostatin production on oncogenicity and metastatic activity of HPV16-transformed mouse cells: Role of interleukin 1α

Two mouse HPV16-transformed cell lines, viz. MK16 cells, which induce metastasizing tumors, and TC-1 cells, which induce non-metastasizing tumors were transduced with the gene for mouse endostatin. Two clones constitutively expressing endostatin were isolated from each of them. They were denoted ME3 and ME9, and TE2 and TE5, respectively. When inoculated into mice, ME3 cells were non-oncogenic. Nearly all mice inoculated with ME9 cells developed tumors, but considerably later than did the parental MK16 cells and metastasis formation was strongly reduced in these animals. On the other hand, TE2 and TE5 cells displayed oncogenic potential similar to that of the parental cells. To provide more information on these different effects of endostatin production, cell lysates of all six lines studied were tested for the content of 25 factors known to be involved in angiogenesis. The parental MK16 cells differed from the parental TC-1 cells and also from all endostatin producing sublines by a markedly higher production of interleukin 1alpha (IL-1alpha) and, to a lesser extent, by a higher production of several other factors tested. Additional experiments indicated that the suppression of the production of IL-1alpha by the parental MK16 caused by endostatin was due to an autocrine mechanism.

Polymorphism of IL-1alpha(-889) Gene and Its Association with Aggressive Periodontitis.

Adult periodontitis is a complex multifactorial disease whose etiology is not well defined. The pro-inflammatory and bone resorption properties of Interleukine-1alpha (IL-1alpha) strongly suggest a role for this cytokine in the pathogenesis of periodontal disease. Eighty Iranian adult patients with periodontitis and 80 Iranian controls were investigated in this study.In this study we report that the frequency of IL-1alpha genotypes including allele 2 of the IL-1alpha(-889) restriction fragment length bi-allele polymorphism were significantly increased in patients with advanced aggressive adult periodontitis compared to those with early and moderate disease. Furthermore, allele 2 was associated with increased production of IL-1alpha by activated peripheral blood polymorphonuclear cells of patients with advanced disease, although this increase failed to reach statistical significance. Finally, the data obtained revealed significant linkage disequilibrium between allele 2 of the IL-1alpha (-889) polymorphism and allele 2 of the bi-allelic IL-1beta (+3953) polymorphism in both patients and orally normal controls. These findings provide new insight into the possible rate of IL-1alpha and beta genes polymorphism in the susceptibility to adult periodontitis.

日本産赤米,黒米,白米のぬか抽出液中に存在する免疫調節活性

To determine the potent immunomodulating activities of different types of Japanese rice bran, we analyzed the effects of extracts of red, brown and black rice brans on the cell proliferation and cytokine production of mouse immunocompetent cells by cell culture experiments. Methanol extracts of these rice brans showed suppressive activities against the proliferative response (3H-thymidine incorporation) of mouse spleen cells induced by concanavalin A (Con A) or lipopolysaccharide (LPS) in the cell culture experiments. Although the black and brown rice bran extracts showed suppressive effects on the production of interferon gamma (IFN gamma) or interleukin 6 (IL-6) in mouse spleen cells induced by Con A or LPS, the red rice bran extract exhibited stimulatory effects on the same cytokine-producing systems. Furthermore, when the effects of these extracts on the production of macropahage-derived inflammatory cytokines such as interleukin-1alpha (IL-1alpha) and tumor necrosis factor alpha (TNF-alpha) were assayed, the red rice bran extract caused a stimulatory effect on the IL-1alpha production from mouse macrophages induced by LPS, but did not show a significant effect on TNF-alpha production. However, the brown and black rice bran extracts exhibited significant inhibitory effects on the production of IL-1alpha and TNF-alpha in the same macrophage culture experiment. A possible mechanism of the immunomodulating activities of the rice bran extracts and the immunopharmacological significance of these findings are discussed.

Effects of Coccoloba uvifera L. on UV‐stimulated melanocytes

Exposure to ultraviolet (UV) radiation induces generation of reactive oxygen species, production of proinflammatory cytokines and melanocyte-stimulating hormone (MSH) as well as increase in tyrosinase activity. The potential photoprotective effects of Coccoloba uvifera extract (CUE) were evaluated in UV-stimulated melanocytes. Human epidermal melanocytes were used as an in vitro model to evaluate the effects of CUE on the production interleukin-1alpha (IL-1alpha), tumor necrosis factor alpha (TNF-alpha), and alpha-MSH under basal and UV-stimulated conditions. Antioxidant and anti-tyrosinase activities were also evaluated in membrane lipid peroxidation and mushroom tyrosinase assay, respectively. Coccoloba uvifera L. showed antioxidant and anti-tyrosinase activities and also inhibited the production of IL-1alpha, TNF-alpha and alpha-MSH in melanocytes subjected to UV radiation (P<0.01). Moreover, CUE inhibited the activity of tyrosine kinase in cell culture under basal and UV radiation conditions (P<0.001), corroborating the findings of the mushroom tyrosinase assay. This study supports the photoprotective potential of CUE.

In vitromodulation of human keratinocyte pro- and anti-inflammatory cytokine production by the capsule ofMalasseziaspecies

Malassezia spp. are commensal, cutaneous fungi that are implicated in seborrhoeic dermatitis. We hypothesize that the lipid-rich capsule of Malassezia spp. masks the organism from host detection, and depletion of this layer elicits an inflammatory response. To test this, preparations of capsulated or acapsular [10% (v/v) Triton X-100 treated], viable and nonviable, exponential or stationary phase Malassezia furfur, Malassezia globosa, Malassezia obtusa, Malassezia restricta, Malassezia slooffiae and Malassezia sympodialis, were incubated with normal human keratinocytes. Proinflammatory (IL-6, IL-8, IL-1alpha and tumour necrosis factor-alpha) and anti-inflammatory cytokine (IL-10) release and intracellular IL-10 concentrations were quantified using enzyme-linked immunosorbent assays. Capsulated Malassezia yeasts stimulated limited or no production of inflammatory cytokines, and increased intracellular IL-10 (P < 0.05). Removal of the capsule of many Malassezia preparations caused a significantly increased production of IL-6, IL-8 and IL-1alpha, and a decrease in intracellular IL-10. Notably, acapsular viable, stationary phase M. globosa caused a 66-fold increase in IL-8 production (P < 0.001) and acapsular nonviable, stationary phase M. furfur caused a 38-fold increase in IL-6 production (P < 0.001) and a 12-fold decrease in intracellular IL-10 (P < 0.001). These results support the hypothesis that the lipid layer of Malassezia spp. modulates cytokine production by keratinocytes. This has implications in the pathogenesis of seborrhoeic dermatitis.