Sodium arsenite impairs insulin secretion and transcription in pancreatic beta-cells

Human studies have shown that chronic inorganic arsenic (iAs) exposure is associated with a high prevalence and incidence of type 2 diabetes. However, the mechanism(s) underlying this effect are not well understood, and practically, there is no information available on the effects of arsenic on pancreatic beta-cells functions. Thus, since insulin secreted by the pancreas plays a crucial role in maintaining glucose homeostasis, our aim was to determine if sodium arsenite impairs insulin secretion and mRNA expression in single adult rat pancreatic beta-cells. Cells were treated with 0.5, 1, 2, 5 and 10 microM sodium arsenite and incubated for 72 and 144 h. The highest dose tested (10 microM) decreased beta-cell viability, by 33% and 83%, respectively. Insulin secretion and mRNA expression were evaluated in the presence of 1 and 5 microM sodium arsenite. Basal insulin secretion, in 5.6 mM glucose, was not significantly affected by 1 or 5 microM treatment for 72 h, but basal secretion was reduced when cells were exposed to 5 microM sodium arsenite for 144 h. On the other hand, insulin secretion in response to 15.6 mM glucose decreased with sodium arsenite in a dose-dependent manner in such a way that cells were no longer able to distinguish between different glucose concentrations. We also showed a significant decrease in insulin mRNA expression of cells exposed to 5 microM sodium arsenite during 72 h. Our data suggest that arsenic may contribute to the development of diabetes mellitus by impairing pancreatic beta-cell functions, particularly insulin synthesis and secretion.     

Inhibition of T lymphocyte activation by estramustine: dose-dependent interference with IL-2 production and later proliferation events

Estramustine, a nitrogen mustard derivative of estradiol-17 beta widely used for treatment of prostatic cancer, was found to inhibit the proliferative response of mouse lymphocytes to T (concanavalin A) and B (lipopolysaccharide) cell mitogens in vitro. Concanavalin A-induced lymphoproliferation was considerably more sensitive to the antiproliferative effect of estramustine than lipopolysaccharide-stimulated proliferation. The concentration of estramustine selectively inhibiting T lymphocyte proliferation was only active when present during the first 24 h of culture and could be overcome by exogenously added interleukin 2. Estramustine was shown to directly inhibit the production of interleukin 2 in concanavalin A-stimulated lymphocyte cultures without affecting the expression of interleukin 2 receptors. Thus the preferential inhibitory effect of estramustine on mitogen-induced T lymphocyte activation is apparently mediated by interference with the production or release of interleukin 2.     

Sodium diethyldithiocarbamate restores T lymphocyte proliferation, interleukin-2 production and NK activity in cyclophosphamide-immunosuppressed animals

The in vivo effect of sodium diethyldithiocarbamate (DTC) on IL-2 production, mitogen-induced proliferation and natural killer (NK) activity of lymphocytes from normal as well as cyclophosphamide (CY)-treated mice has been investigated. DTC was given in a single subcutaneous injection (25 mg/kg) to normal or cyclophosphamide-treated mice (250 mg/kg i.p. simultaneously to DTC). An enhancement of T lymphocyte proliferation in CY-treated animals and an increase of IL-2 production in both normal and immunosuppressed mice was observed. When DTC (10 mg/kg) was administered daily for two weeks an increase in concanavalin A-induced mitogenesis, IL-2 production and NK activity in CY-treated animals was observed. These immune parameters were reduced 12 days after CY treatment by a factor of 2 to 3 times, while DTC treatment restored these responses to normal levels. LPS-induced mitogenesis was not significantly enhanced. The effect of DTC could be partially mediated by changes in IL-2 activity. According to these results some functional parameters of the immune system of the suppressed host can be partially or completely restored by means of an appropriate immunomodulator treatment. DTC could be of interest in the treatment of diseases where immune functions are impaired or in combined treatments with immunosuppressants.     

Age-related effects of sodium arsenite on splenocyte proliferation and Th1/Th2 cytokine production

Aging is associated with immune dysfunction and conditions such as inflamm-aging and immunosuppression. Arsenic, an environmental contaminant distributed worldwide, affects the immune system. This study tested the hypothesis that arsenic has distinct effects on T cell proliferation and the production of cytokines by activated T cells. Murine splenocytes from young (2 months) and aged (24–26 months) C57BL/6 mice were exposed to arsenite (As³⁺), the most toxic form of inorganic arsenic, and stimulated with concanavalin A (Con A) or anti-CD3 antibody. T cell proliferation decreased significantly in response to Con A and anti-CD3 at subtoxic doses of arsenite in splenocytes from both young and aged mice. Arsenite, added concurrently with Con A or anti-CD3, significantly inhibited the production of interleukin-2 (IL-2), interferon-γ (IFN-γ), and interleukin-4 (IL-4) by splenocytes from young mice and significantly reduced the production of IL-10 by splenocytes from aged mice. In contrast, the production of IL-2 and IL-4 by splenocytes from aged mice was only slightly affected by arsenite. The results show that arsenic exposure reduces the immune response in splenocytes. Moreover, this effect may be influenced by aging.     

Sodium arsenite inhibits migration of extravillous trophoblast cells in vitro

This study used a first-trimester human extravillous trophoblast (EVT) cell line, HTR-8/SVneo, to investigate whether sodium arsenite (AsNaO₂) reduces human EVT migration and invasion. Treatments with 2.5 μM AsNaO₂ or less (≤187.3 μg/L), concentrations that are relevant to human exposures in drinking water, were sublethal to HTR-8/SVneo cells. A 72-h exposure to sodium arsenite inhibited cell migration in a concentration-dependent manner at 0.625, 1.25 and 2.5 μM. Significant changes in cell proliferation were not observed under these treatment conditions. Moreover, inhibition of cell migration was unrelated to phosphorylation of focal adhesion kinase Tyr397. In contrast to cell migration, 72-h exposures to AsNaO₂ (0.3125–2.5 μM) had no significant effects on cell invasion, nor on the activities and protein expression of matrix metalloproteinase (MMP) 2 and MMP9. Because trophoblast migration is important for placentation, these results suggest an effect that could contribute to insufficiency of placental development and adverse pregnancy outcomes.     

Sodium arsenite inhibits terminal differentiation of murine C3H 10T1/2 preadipocytes

Cancer represents an imbalance between cell proliferation and differentiation, two processes that are coordinately and antagonistically regulated. Aberrant cell proliferation is considered to be an important etiological factor in the development of arsenic-induced cancer, suggesting that arsenic also dysregulates differentiation. Based on evidence that arsenic modulates mitogenic events that antagonize the process of differentiation, this study addresses the hypothesis that sodium arsenite inhibits insulin/dexamethasone-induced differentiation of C3H 10T1/2 preadipocytes; it was further postulated that arsenic-treated cells retain mitogenic responsiveness under differentiating conditions. To test this hypothesis, the differentiation capacity of C3H 10T1/2 preadipocytes was examined in control cells and cells treated with sodium arsenite. Differentiation was assessed morphologically and quantified by Oil Red-O staining of accumulated lipids. The effect of long-term arsenic exposure on mitogenic competence was quantified by flow cytometry, [(3)H]thymidine incorporation, and cell counting under conditions favorable for adipocyte differentiation. Results indicate that arsenic inhibits morphological differentiation of wild-type C3H 10T1/2 preadipocytes. Short-term arsenic exposure inhibits differentiation in a dose-dependent manner, with arsenic concentrations > or = 3 microM producing a significant inhibition of dexamethasone/insulin-induced lipid accumulation. Furthermore, arsenic-treated cells exhibit an accentuated response to mitogenic stimulation under differentiating conditions. These data suggest that arsenic exposure results in the inhibition of cellular programming required for terminal differentiation of C3H 10T1/2 preadipocytes and that cells acquire mitogenic hyperresponsiveness. The ability of arsenic to dysregulate the balance between proliferation and differentiation is proposed to be one mechanism by which this metalloid causes cancer in humans.     

Recombinant human B7-H4 expressed in Escherichia coli inhibits T lymphocyte proliferation and IL-2 secretion in vitro

AIM: To explore the biofunctions of human B7-H4 generated from prokaryotic system. METHODS: The gene of human B7-H4 extracellular region (IgV-like and IgC-like domains) was obtained by PCR from human cDNA FLJ22418 and then inserted into the prokaryotic expression vector pGEX-5X-3 expressing glutathione s-transferase (GST) fusion protein. After being identified by restriction enzyme digestion and sequencing, the recombinant vector was transferred into host strain E coli BL21-RIL(DE3). A 47 kDa fusion protein (GST/hB7-H4) was induced by isopropyl-beta-D-thiogalactopyranoside (IPTG) and purified by standard methods reported in the prokaryotic system. The inhibitory effect of GST/hB7-H4 on proliferation of T cells was observed in vitro by CD3mAb activated T-cell culturing system and [(3)H]-thymidine incorporation assay. The concentrations of interleukin-2 and iterferon-g in the supernatants of T cells were determined by ELISA. RESULTS: We successfully constructed the method for high-level expression and purification of the hB7-H4 extracellular domain as GST fusion protein from E coli. The GST/hB7-H4 fusion protein produced in bacteria had obvious biological activity to inhibit T-lymphocyte proliferation and IL-2 secretion. CONCLUSION: The prokaryote expression system could be used to generate hB7-H4 protein with natural spatial conformations and biological functions, which provided an efficient and economical way for the preparation of this target protein.     

丁酸钠调控HepG2细胞的增殖、凋亡和侵袭

目的 研究丁酸钠对人肝癌细胞HepG2增殖和凋亡的影响,并探讨可能的作用机制。方法 用不同浓度丁酸钠处理HepG2 细胞后,MTT方法检测细胞的增殖能力,流式细胞技术检测细胞周期的分布和细胞凋亡,Transwell小室检测丁酸钠对HepG2细胞侵袭能力的影响。免疫荧光法检测HDAC4蛋白在HepG2细胞中的表达及定位。蛋白质印迹法(Western Blot)检测HDAC4蛋白的表达水平。结果 随着丁酸钠处理浓度的增加和处理时间的延长,HepG2细胞增殖能力明显受抑制,细胞周期也发生阻滞,G1期细胞比例明显增加,而S期细胞比例明显减少。不同浓度(0,1,5,10 mmol/L)丁酸钠处理HepG2细胞24 h后,早期凋亡率分别为2.7％,4.5％,6.5％,6.7%,差异有统计学意义(F=15.1,P=0.001)。丁酸钠显著抑制细胞侵袭能力,侵袭细胞百分比分别降至72.7%(1 mmol/L)、41.7%(5 mmol/L)、21.3%(10 mmol/L),差异有统计学意义(F=202.1,P<0.001)。HDAC4蛋白在HepG2 细胞中呈阳性表达,主要位于细胞质中。丁酸钠明显抑制HDAC4蛋白的表达,并呈浓度依赖性。结论 丁酸钠抑制肝癌细胞系HepG2的增殖,调控细胞周期、促进凋亡,抑制细胞的侵袭能力。     

In vitro ozone exposure inhibits mitogen-induced lymphocyte proliferation and IL-2 production

Human blood mononuclear cells were exposed to ozone in vitro and thereafter analyzed for competence in mitogen-induced proliferation as well as IL-1 and IL-2 production. Proliferative responses induced by phytohemagglutinin (PHA), concanavalin A (Con A), and pokeweed mitogen (PWM) were all depressed in lymphocytes exposed to an ozone concentration of 1 ppm for 4-6 h. The response to PWM was most sensitive to the ozone effect (38% suppression); responses to Con A and PHA were suppressed to a lesser extent, 23% and 18%, respectively, and were not significantly different from each other. PWM responses were affected at an ozone concentration as low as 0.1 ppm; however, no suppression of Con A-induced proliferation was seen below 0.18 ppm or of PHA-induced proliferation below 0.5 ppm. When lymphocytes and monocytes were exposed separately to ozone and then mixed back with control air-exposed monocytes or lymphocytes, both cell types appeared to be affected and the functional defects caused by the pollutant were additive. Monocyte IL-1 production induced by endotoxin was not affected by ozone exposure, while surface expression of HLA-DR on exposed monocytes was reduced by 40% 24 h after exposure. Moreover, lymphocytes exposed to ozone produced 46% less IL-2 while expressing similar surface density of IL-2 receptors. Taken together, these results show that exposure to ozone has distinct adverse effects on lymphocytes and monocytes, both of which are important in local immune defenses in the lung.     

褪黑素对海洛因依赖大鼠淋巴细胞增殖和IL-2产生的影响

目的　观察褪黑素 (MT)对海洛因 (Her)依赖大鼠淋巴细胞增殖和IL 2产生的影响。方法　以剂量递增连续scHer建立海洛因大鼠依赖模型 ,并同时随机设一组给予MT保护。 42d后随机分组 ,设MT治疗组、美沙酮治疗组、自然戒断组、依赖组 ;另选正常大鼠为溶媒对照组。采用MTT法观察MT对ConA诱导的大鼠脾淋巴细胞增殖反应和IL 2的产生。结果　MT保护组 37 5mg·kg-1,bid与依赖组比较 ,对ConA诱导的T淋巴细胞增殖反应和IL 2的产生有明显的促进作用 ;MT 12 5、37 5、6 2 5mg·kg-1,bid 3个剂量组与美沙酮治疗组、自然戒断组比较 ,对ConA诱导的T淋巴细胞增殖反应和IL 2的产生均有不同程度的促进作用。结论　MT对海洛因造成的细胞免疫功能低下可能有预防和逆转的作用。     

落地生根对小鼠免疫功能的影响

目的 :探讨中药落地生根对小鼠免疫功能的影响。方法 :用落地生根水浸出液给小鼠灌胃 ,检测小鼠脾淋巴细胞增殖和IL 2的产生及骨髓细胞的增殖反应。结果 :高剂量落地生根 (8mg·kg-1,ig) ,能显著增强脾淋巴细胞增殖反应 ,与生理盐水对照组比较有非常显著性差异 (P <0 .0 1 ) ;对细胞因子IL 2的产生也有促进作用 ,与生理盐水对照组比较有显著性差异(P <0 .0 5 )。但对骨髓细胞增殖无明显影响 ,低剂量落地生根 (5g·kg-1,ig)对以上指标均无明显影响。结论 :落地生根可以增强小鼠的免疫功能     

Echinacea alkylamides inhibit interleukin-2 production by Jurkat T cells

Alkylamides present in Echinacea species have reported immunomodulatory actions, yet their direct effects on T lymphocytes, key mediators of antiviral immunity, are poorly understood. We hypothesized that constituents present in ethanolic extracts of Echinacea species exert direct immunomodulatory effects on human Jurkat T cells. Modulation of IL-2 production by submaximally stimulated Jurkat cells was determined in response to treatment with extracts prepared from dried aerial parts of Echinacea purpurea. Cells were treated with the extracts, with alkylamides or caffeic acid derivatives isolated from Echinacea species, or with corresponding ethanol vehicle, in the absence or presence of phytohemagglutinin and phorbal ester. E. purpurea extracted in a solvent mixture of 95:5 ethanol/water dose-dependently inhibited IL-2 production. This IL-2 inhibitory activity correlated with the presence of alkylamides but not caffeic acid derivatives, as determined by high performance liquid chromatography/electrospray ionization-mass spectrometry analysis. Simultaneous measurement of secreted IL-2 by ELISA and cell viability by the XTT assay showed that the 95:5 ethanol/water extract of E. purpurea was both IL-2 suppressive and cytotoxic at 50 and 100 microg/mL. Lower concentrations from 6.25 to 25 microg/mL significantly decreased IL-2 production but not cell viability. Alkylamides at concentrations found in a 50 microg/mL extract decreased IL-2 production by approximately 50%. Two Echinacea-derived alkylamides significantly depressed IL-2 production but not cell viability in a dose-dependent manner. Thus, alkylamides present in E. purpurea suppress the ability of activated Jurkat T cells to produce IL-2 independently of direct, cytotoxic effects.     

Regulatory mechanisms of IL-2 and IFNgamma suppression by quercetin in T helper cells

Quercetin is a popular flavonoid compound that is biosynthesized by plants; it is suggested to modulate a variety of inflammatory responses of macrophages and T lymphocytes. Oral administration of quercetin in arthritic rats dramatically diminishes clinical signs of arthritis. Moreover, quercetin ameliorates experimental autoimmune encephalomyelitis, which is associated with Th1-mediated immune responses. Like quercetin inhibits macrophage-induced cytokine production, it also blocks IL-12-dependent JAK-STAT signaling in Th cells. Despite the anti-inflammatory effects of quercetin acting through Th cells, the regulatory mechanisms remain unclear. Here, we studied the function of quercetin in Th cells and found that quercetin suppressed both IFNgamma and IL-2 production upon T cell receptor stimulation. Furthermore, we uncovered the regulatory mechanisms of quercetin involved in the inhibition of cytokine production during Th cell activation. The fact that quercetin-derived IFNgamma suppression was blocked in T-bet-deficient Th cells demonstrated quercetin act through the modulation of T-bet expression. Whereas IL-2 inhibition by quercetin was independent of T-bet expression, quercetin diminished IL-2R alpha expression, which is critical for positive regulatory loop of IL-2 autoactivation. Taken together, quercetin is suggested to repress both IFNgamma and IL-2 cytokine production by independent mechanisms; T-bet-dependent IFNgamma suppression and IL-2R alpha-dependent IL-2 inhibition.     

Differential influences of the BPA,BPS and BPF on in vitro IL-17 secretion by mouse and human T cells

The endocrine disruptor and food contaminant bisphenol A (BPA) is frequently present in consumer plastics and can produce several adverse health effects participating in the development of inflammatory and autoimmune diseases. Regulatory restrictions have been established to prevent risks for human health, leading to the substitution of BPA by structural analogues, such as bisphenol S (BPS) and F (BPF). In this study, we aimed at comparing the in vitro impact of these bisphenols from 0.05 to 50,000 nM on Th17 differentiation, frequency and function in mouse systemic and intestinal immune T cells and in human blood T cells. This study reports the ability of these bisphenols, at low and environmentally relevant concentration, i.e, 0.05 nM, to increase significantly IL-17 production in mouse T cells but not in human T lymphocytes. The use of an aryl hydrocarbon receptor (AhR) specific inhibitor demonstrated its involvement in this bisphenol-induced IL-17 production. We also observed an increased IL-17 secretion by BPS and BPF, and not by BPA, in mouse naive T cells undergoing in vitro Th17 differentiation. In total, this study emphasizes the link between bisphenol exposures and the susceptibility to develop immune diseases, questioning thus the rational of their use to replace BPA.     

Etidronate inhibits the production of IL-6 by osteoblast-like cells

IL-6 is a resorbing cytokine synthesized by osteoblasts and monocytes that has been implicated in the pathogenesis of osteoporosis. Bisphosphonates are well-known antiresorptive drugs, the antiosteoclastic effect of which has been recently suggested to be brought about at least in part through osteoblasts. Based on these facts, we have studied the effect of etidronate on the production of IL-6 by two tumoral cell lines of human osteoblastic phenotype (MG63 and SaOs cells), and by peripheral blood mononuclear cells (PBMC). For comparison, another antiresorptive drug, estradiol, was included in the study. MG63 cells were stimulated with LPS and IL-1 beta, SaOs cells with LPS, IL-1 beta and PMA, and PBMC with LPS and PMA. Etidronate was tested at 10(-7), 10(-6), 10(-5), and 10(-4) M, and 17beta-estradiol was tested at 10(-10), 10(-9), 10(-8), and 10(-7) M. IL-6 was determined in supernatants by an ELISA. No significant effect of either etidronate or estradiol on IL-6 secretion by LPS or PMA-stimulated PBMC was found. However, in osteoblastic-like cells, an inhibition of IL-6 production by etidronate in LPS-stimulated cultures was found. At the highest concentrations tested, IL-6 production values were 58 +/- 9% and 53 +/- 8% of those at base line for MG63 and SaOs cells, respectively. Estradiol did not modify IL-6 secretion under any condition. In conclusion, our study supports the contention that the antiresorptive effect of bisphosphonates may be due in part to a decrease in IL-6 production by osteoblasts.     

Chalcones from Chinese liquorice inhibit proliferation of T cells and production of cytokines

Licochalcone A (LicA), an oxygenated chalcone, has been shown to inhibit the growth of both parasites and bacteria. In this study, we investigated the effect of LicA and four synthetic analogues on the activity of human peripheral blood mononuclear cell proliferation and cytokine production. Four out of five chalcones tested inhibited the proliferation of lymphocytes measured by thymidine incorporation and by flow cytometry. The production of pro- and anti-inflammatory cytokines from monocytes and T cells was also inhibited by four of five chalcones. Furthermore, intracellular detection of cytokines revealed that the chalcones inhibited the production rather than the release of the cytokines. Taken together, these results indicate that LicA and some analogues may have immunomodulatory effects, and may thus be candidates not only as anti-microbial agents, but also for the treatment of other types of diseases.