HPV16 Tumor Associated Macrophages Suppress Antitumor T Cell Responses

Purpose: High-risk human papillomavirus (HPV) is the main etiologic factor for cervical cancer. The severity of HPV-associated cervical lesions has been correlated to the number of infiltrating macrophages. The objective of this work is to characterize the role of tumor-associated macrophages (TAM) on the immune cellular response against the tumor. Experimental Design: We used the HPV16 E6- and E7-expressing TC-1 mouse tumor model to study the effect of TAM on T-cell function in vitro, and depleted TAM, using clodronate-containing liposomes, to characterize its role in vivo. Results: TAM, characterized by the positive expression of CD45, F4/80, and CD11b, formed the major population of infiltrating tumor cells. TAM displayed high basal Arginase I activity, producing interleukin-10 (IL-10); they were resistant to iNOSll activity induction, therefore reversion to M1 phenotype, when stimulated in vitro with lipopolysaccharide/IFN gamma, indicating an M2 phentoype. In cultures of isolated TAM, TAM induced regulatory phenotype, characterized by IL-10 and Foxp3 expression, and inhibited proliferation of CD8 lymphocytes. In vivo, depletion of TAM inhibited tumor growth and stimulated the infiltration of tumors by HPV16 E7(49-57)-specific CD8 lymphocytes, whereas depletion of Gr1(+) tumor-associated cells had no effect. Conclusions: M2-like macrophages infiltrate HPV16-associated tumors causing suppression of antitumor T-cell response, thus facilitating tumor growth. Depletion or phenotype alteration of this population should be considered in immunotherapy strategies.

Structural changes in the epithelium of the small intestine and immune cell infiltration of enteric ganglia following acute mucosal damage and local inflammation

An acute enteritis is commonly followed by intestinal neuromuscular dysfunction, including prolonged hyperexcitability of enteric neurons. Such motility disorders are associated with maintained increases in immune cells adjacent to enteric ganglia and in the mucosa. However, whether the commonly used animal model, trinitrobenzene sulphonate (TNBS)-induced enteritis, causes histological and immune cell changes similar to human enteric neuropathies is not clear. We have made a detailed study of the mucosal damage and repair and immune cell invasion following intralumenal administration of TNBS. Intestines from untreated, sham-operated and TNBS-treated animals were examined at 3 h to 56 days. At 3 h, the mucosal surface was completely ablated, by 6 h an epithelial covering was substantially restored and by 1 day there was full re-epithelialisation. The lumenal epithelium developed from a squamous cell covering to a fully differentiated columnar epithelium with mature villi at about 7 days. Prominent phagocytic activity of enterocytes occurred at 1-7 days. A surge of eosinophils and T lymphocytes associated with the enteric nerve ganglia occurred at 3 h to 3 days. However, elevated immune cell numbers occurred in the lamina propria of the mucosa until 56 days, when eosinophils were still three times normal. We conclude that the disruption of the mucosal surface that causes TNBS-induced ileitis is brief, a little more than 6 h, and causes a transient immune cell surge adjacent to enteric ganglia. This is much briefer than the enteric neuropathy that ensues. Ongoing mucosal inflammatory reaction may contribute to the persistence of enteric neuropathy.

The Impact of Vitamin A Supplementation on the Immune System of Vitamin A-deficient Children

Background & Aims: To investigate the effect of vitamin A supplementation on parameters of the immune system of vitamin A-deficient children. Methods: The study was carried out in four phases: 1) determination of serum retinol in 631 children from 36 to 83 months of age; 2) assessment of immunological markers [immunoglobulins and complement fractions, immunophenotyping of T and B lymphocytes, and natural killer (NK) cells], blood count, and serum ferritin of 52 vitamin A-deficient children (serum retinol <0.70 mu mol/L); 3) supplementation of the 52 deficient children with 200,000 IU of vitamin A; 4) determination of serum retinol and the immunological parameters 2 months after vitamin A supplementation. Results: Before vitamin A supplementation, 24.0% of the children were anemic and 4.3 %had reduced ferritin concentrations. There was no significant difference between mean values of retinol according to the presence/absence of anemia. The mean values of the humoral and cellular immunological parameters did not show a statistically significant difference before and after supplementation with vitamin A. Children with concomitant hypovitaminosis A and anemia presented a significant increase in absolute CD4 and CD8 T-cell counts after vitamin A supplementation (p < 0.05). Conclusion: Vitamin A had an effect on the recruitment of T and B lymphocytes to the circulation of children with hypovitaminosis A and anemia.

Immunomodulatory effects of Pteridium aquilinum on natural killer cell activity and select aspects of the cellular immune response of mice

Pteridium aquilinum (bracken fern) is one of the most common plants. Epidemiological studies have revealed a higher risk of certain types of cancers (i.e., esophageal, gastric) in people who consume bracken fern directly ( as crosiers or rhizomes) or indirectly through the consumption of milk from livestock that fed on the plant. In animals, evidence exists regarding the associations between chronic bracken fern intoxication, papilloma virus infection, and the development of carcinomas. While it is possible that some carcinogens in bracken fern could be responsible for these cancers in both humans and animals, it is equally plausible that the observed increases in cancers could be related to induction of an overall immunosuppression by the plant/its various constituents. Under the latter scenario, normal tumor surveillance responses against nascent (non-bracken-induced) cancers or responses against viral infections ( specifically those linked to induction of cancers) might be adversely impacted by continuous dietary exposure to this plant. Therefore, the overall objective of this study was to evaluate the immunomodulatory effects of bracken fern following daily ingestion of its extract by a murine host over a period of 14 ( or up to 30) days. In C57BL/6 mice administered ( by gavage) the extract, histological analyses revealed a significant reduction in splenic white pulp area. Among a variety of immune response parameters/functions assessed in these hosts and isolated cells, both delayed-type hypersensitivity (DTH) analysis and evaluation of IFN gamma. production by NK cells during T(H)1 priming were also reduced. Lastly, the innate response in these hosts-assessed by analysis of NK cell cytotoxic functionality-was also diminished. The results here clearly showed the immunosuppressive effects of P. aquilinum and that many of the functions that were modulated could contribute to the increased risk of cancer formation in exposed hosts.

Effects of exercise training on autonomic dysfunction management in an experimental model of menopause and myocardial infarction

Objective: The aim of this study was to investigate the effects of exercise training on cardiovascular autonomic dysfunction in ovariectomized rats submitted to myocardial infarction. Methods: Female Wistar rats were divided into the following ovariectomized groups: sedentary ovariectomized (SO), trained ovariectomized (TO), sedentary ovariectomized infarcted (SOI), and trained ovariectomized infarcted (TOI). Trained groups were submitted to an exercise training protocol on a treadmill (8 wk). Arterial baroreflex sensitivity was evaluated by heart rate responses to arterial pressure changes, and cardiopulmonary baroreflex sensitivity was tested by bradycardic and hypotension responses to serotonin injection. Vagal and sympathetic effects were calculated by pharmacological blockade. Results: Arterial pressure was reduced in the TO in comparison with the SO group and increased in the TOI in relation to the SOI group. Exercise training improved the baroreflex sensitivity in both the TO and TOI groups. The TOI group displayed improvement in cardiopulmonary reflex sensitivity compared with the SOI group at the 16 mu g/kg serotonin dose. Exercise training enhanced the vagal effect in both the TO (45%) and TOI (46%) animals compared with the SO and SOI animals and reduced the sympathetic effect in the TOI (38%) in comparison with the SOI animals. Significant correlations were obtained between bradycardic baroreflex responses and vagal (r = -0.7, P < 0.005) and sympathetic (r = 0.7, P < 0.001) effects. Conclusions: These results indicate that exercise training in ovariectomized rats submitted to myocardial infarction improves resting hemodynamic status and reflex control of the circulation, which may be due to an increase in the vagal component. This suggests a homeostatic role for exercise training in reducing the autonomic impairment of myocardial infarction in postmenopausal women.

Lifetime Overproduction of Circulating Angiotensin-(1-7) Attenuates Deoxycorticosterone Acetate-Salt Hypertension-Induced Cardiac Dysfunction and Remodeling

We evaluated the development of arterial hypertension, cardiac function, and collagen deposition, as well as the level of components of the renin-angiotensin system in the heart of transgenic rats that overexpress an angiotensin (Ang)-(1-7)-producing fusion protein, TGR(A1-7)3292 (TG), which induces a lifetime increase in circulating levels of this peptide. After 30 days of the induction of the deoxycorticosterone acetate (DOCA)-salt hypertension model, DOCA-TG rats were hypertensive but presented a lower systolic arterial pressure in comparison with DOCA-Sprague-Dawley (SD) rats. In contrast to DOCA-SD rats that presented left ventricle (LV) hypertrophy and diastolic dysfunction, DOCA-TG rats did not develop cardiac hypertrophy or changes in ventricular function. In addition, DOCA-TG rats showed attenuation in mRNA expression for collagen type I and III compared with the increased levels of DOCA-SD rats. Ang II plasma and LV levels were reduced in SD and TG hypertensive rats in comparison with normotensive animals. DOCA-TG rats presented a reduction in plasma Ang-(1-7) levels; however, there was a great increase in Ang-(1-7) (approximate to 3-fold) accompanied by a decrease in mRNA expression of both angiotensin-converting enzyme and angiotensin-converting enzyme 2 in the LV. The mRNA expression of Mas and Ang II type 1 receptors in the LV was not significantly changed in DOCA-SD or DOCA-TG rats. This study showed that TG rats with increased circulating levels of Ang-(1-7) are protected against cardiac dysfunction and fibrosis and also present an attenuated increase in blood pressure after DOCA-salt hypertension. In addition, DOCA-TG rats showed an important local increase in Ang-(1-7) levels in the LV, which might have contributed to the attenuation of cardiac dysfunction and prefibrotic lesions. (Hypertension. 2010;55:889-896.)

Homocysteine Thiolactone Induces Cardiac Dysfunction: Role of Oxidative Stress

This study investigates the cardiac functioning in male Wistar rats after treatments with methionine and homocysteine thiolactone (HcyT). The rats were distributed into 3 groups and treated for 8 weeks. Group I was the control (CO) group, given water, group II was treated with methionine, and group III with HcyT (100 mg/kg). Morphometric and functional cardiac parameters were evaluated by echocardiography. Superoxide dismutase (SOD), catalase, and glutathione S-transferase activities, chemiluminescence, thiobarbituric acid reactive substances, and immunocontent were measured in the myocardium. Hyperhomocysteinemia was observed in rats submitted to the both treatments. The results showed diastolic function was compromised in HcyT group, seen by the increase of E/A (peak velocity of early (E) and late (A) diastolic filling) ratio, decrease in deceleration time of E wave and left ventricular isovolumic relaxation time. Myocardial performance index was increased in HcyT group and was found associated with increased SOD immunocontent. HcyT group demonstrated an increase in SOD, catalase, and glutatione S-transferase activity, and chemiluminescence and thiobarbituric acid reactive substances. Overall, these results indicated that HcyT induces a cardiac dysfunction and could be associated with oxidative stress increase in the myocardium.

Reversal of Postprandial Endothelial Dysfunction by Cyclooxygenase Inhibition in Healthy Volunteers

The aim of this study was to evaluate the role of cyclooxygenase (COX) in venous vascular reactivity changes after an oral lipid overload (OLO). Venous endothelial function (dorsal hand vein technique) was evaluated in fasting, 30 minutes after COX inhibition (aspirin-fasting), 2 to 4 hours after an OLO (1000 kcal, 58% fat), and again after COX inhibition (aspirin-OLO, 600 mg/200 mL water) in 10 healthy adults (age, 28.1 +/- 1.3 years; body mass index, 22.3 +/- 0.6 kg/m(2)). Fasting, 2- to 4-hour post-OLO, and 60-minute postaspirin plasma glucose, insulin, and lipids were also evaluated. The OLO increased triglycerides and insulin, reduced low-density lipoprotein and high-density lipoprotein, but glycemia and total cholesterol remained unchanged. There were no metabolic differences between OLO and aspirin-OLO. In fasting, aspirin reduced acetylcholine-induced venodilation (107.0% +/- 14% versus 57.3% +/- 11%; P < 0.001). Vascular reactivity was blunted after the OLO (phenylephrine dose: 0.3 +/- 0.2 fasting versus 1.9 +/- 0.8 nmol/min after OLO; P < 0.001) and was partially corrected by aspirin (0.4 +/- 0.2; P < 0.001). Similar changes were observed in maximum venodilation after acetylcholine (107.0% +/- 14% fasting versus 60.4% +/- 9% after OLO, P < 0.001; aspirin-OLO: 95.9% +/- 6%; P < 0.001). The responses to sodium nitroprusside remained unchanged during the study. We conclude that the OLO reduction in the endothelium-dependent venoconstruction and venodilation is partially the result of the action of COX.

The Immune-Pineal Axis Stress as a Modulator of Pineal Gland Function

The temporal organization of mammals presents a daily adjustment to the environmental light/dark cycle. The environmental light detected by the retina adjusts the central clock in the suprachiasmatic nuclei, which innervate the pineal gland through a polysynaptic pathway. During the night, this gland produces and releases the nocturnal hormone melatonin, which circulates throughout the whole body and adjusts several bodily functions according to the existence and duration of darkness. We have previously shown that during the time frame of an inflammatory response, pro-inflammatory cytokines, such as tumor necrosis factor-a, inhibit while anti-inflammatory mediators, such as glucocorticoids, enhance the synthesis of melatonin, interfering in the daily adjustment of the light/dark cycle. Therefore, injury disconnects the organism from environmental cycling, while recovery restores the light/dark information to the whole organism. Here, we extend these observations by evaluating the effect of a mild restraint stress, which did not induce macroscopic gastric lesions. After 2 h of restraint, there was an increase in circulating corticosterone, indicating activation of the hypothalamus-pituitary-adrenal (HPA) axis. In parallel, an increase in melatonin production was observed. Taking into account the data obtained with models of inflammation and stress, we reinforce the hypothesis that the activity of the pineal gland is modulated by the state of the immune system and the HPA axis, implicating the darkness hormone melatonin as a modulator of defense responses.

Relation of pretreatment sequence diversity in NS5A region of HCV genotype 1 with immune response between pegylated-INF/ribavirin therapy outcomes

Hepatitis C virus (HCV) infection frequently persists despite substantial virus-specific immune responses and the combination of pegylated interferon (INF)-alpha and ribavirin therapy. Major histocompatibility complex class I restricted CD8+ T cells are responsible for the control of viraemia in HCV infection, and several studies suggest protection against viral infection associated with specific HLAs. The reason for low rates of sustained viral response (SVR) in HCV patients remains unknown. Escape mutations in response to cytotoxic T lymphocyte are widely described; however, its influence in the treatment outcome is ill understood. Here, we investigate the differences in CD8 epitopes frequencies from the Los Alamos database between groups of patients that showed distinct response to pegylated alpha-INF with ribavirin therapy and test evidence of natural selection on the virus in those who failed treatment, using five maximum likelihood evolutionary models from PAML package. The group of sustained virological responders showed three epitopes with frequencies higher than Non-responders group, all had statistical support, and we observed evidence of selection pressure in the last group. No escape mutation was observed. Interestingly, the epitope VLSDFKTWL was 100% conserved in SVR group. These results suggest that the response to treatment can be explained by the increase in immune pressure, induced by interferon therapy, and the presence of those epitopes may represent an important factor in determining the outcome of therapy.

Dehydroepiandrosterone protects against oxidative stress-induced endothelial dysfunction in ovariectomized rats

Cardiovascular disease is less frequent in premenopausal women than in age-matched men or postmenopausal women. Moreover, the marked age-related decline in serum dehydroepiandrosterone (DHEA) level has been associated to cardiovascular disease. The aim of this study was to evaluate the effects of DHEA treatment on vascular function in ovariectomized rats. At 8 weeks of age, female Wistar rats were ovariectomized (OVX) or sham (SHAM) operated and 8 weeks after surgery both groups were treated with vehicle or DHEA (10 mg kg-1 week-1) for 3 weeks. Aortic rings were used to evaluate the vasoconstrictor response to phenylephrine (PHE) and the relaxation responses to acetylcholine (ACh) and sodium nitroprusside (SNP). Tissue reactive oxygen species (ROS) production and SOD, NADPH oxidase and eNOS protein expression were analysed. PHE-induced contraction was increased in aortic rings from OVX compared to SHAM, associated with a reduction in NO bioavailability. Furthermore, the relaxation induced by ACh was reduced in arteries from OVX, while SNP relaxation did not change. The incubation of aortic rings with SOD or apocynin restored the enhanced PHE-contraction and the impaired ACh-relaxation only in OVX. DHEA treatment corrected the increased PHE contraction and the impaired ACh-induced relaxation observed in OVX by an increment in NO bioavailability and decrease in ROS production. Besides, DHEA treatment restores the reduced Cu/Zn-SOD protein expression and eNOS phosphorylation and the increased NADPH oxidase protein expression in the aorta of OVX rats. The present results suggest an important action of DHEA, improving endothelial function in OVX rats by acting as an antioxidant and enhancing the NO bioavailability.

Aldosterone induces endothelial dysfunction in resistance arteries from normotensive and hypertensive rats by increasing thromboxane A(2) and prostacyclin

Background and purpose: The present study was designed to assess whether cyclooxygenase-2 (COX-2) activation is involved in the effects of chronic aldosterone treatment on endothelial function of mesenteric resistance arteries (MRA) from Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR). Experimental approach: Relaxation to acetylcholine was measured in MRA from both untreated and aldosterone-treated strains. Vasomotor responses to prostacyclin and U46619 were also analysed. Release of 6-oxo-prostaglandin (PG)F(1 alpha) and thromboxane B(2) (TxB(2)) was determined by enzyme immunoassay. COX-2 protein expression was measured by western blot. Key results: Aldosterone reduced acetylcholine relaxation in MRA from both strains. In MRA from both aldosterone-treated strains the COX-1/2 or COX-2 inhibitor (indomethacin and NS-398, respectively), Tx2 synthesis inhibitor (furegrelate), prostacyclin synthesis inhibitor (tranylcypromine) or Tx2/PG2 receptor antagonist (SQ 29 548), but not COX-1 inhibitor SC-560, increased acetylcholine relaxation. In untreated rats this response was increased only in SHR. Prostacyclin elicited a biphasic vasomotor response: lower concentrations elicited relaxation, whereas higher concentrations elicited contraction that was reduced by SQ 29 548. Aldosterone increased the acetylcholine-stimulated production of 6-oxo-PGF(1 alpha) and TxB(2) in MRA from both strains. COX-2 expression was higher in both strains of rats treated with aldosterone. Conclusions and implications: Chronic treatment with aldosterone impaired endothelial function in MRA under normotensive and hypertensive conditions by increasing COX-2-derived prostacyclin and thromboxane A(2). As endothelial dysfunction participates in the pathogenesis of many cardiovascular disorders we hypothesize that anti-inflammatory drugs, specifically COX-2 inhibitors, could ameliorate vascular damage in patients with elevated aldosterone production.

Amino acids and diabetes: implications for endocrine, metabolic and immune function

Aberrant alterations in glucose and lipid concentrations and their pathways of metabolism are a hallmark of diabetes. However, much less is known about alterations in concentrations of amino acids and their pathways of metabolism in diabetes. In this review we have attempted to highlight, integrate and discuss common alterations in amino acid metabolism in a wide variety of cells and tissues and relate these changes to alterations in endocrine, physiologic and immune function in diabetes.

Oxidative stress and inflammatory mediators contribute to endothelial dysfunction in high-fat diet-induced obesity in mice

Objective We investigated the effects of high-fat diet-induced obesity on vascular proinflammatory factors and oxidative stress on endothelium-dependent relaxation of the aorta. Methods Female Swiss mice were submitted to a high-fat diet for 16 weeks. At the end of the experimental period, we evaluated blood pressure, relaxation in response to acetylcholine in aortic rings in the absence and the presence of the superoxide anion scavenger, superoxide dismutase (SOD, 150 U/ml), and the nuclear factor (NF)-kappa B inhibitor, sodium salicylate (5 mmol/l). Aortic protein expression of endothelial nitric oxide synthase, Cu/Zn-SOD, NF-kappa B, I kappa B-alpha, and proinflammatory cytokines were also evaluated. Results Obese mice presented higher systolic and diastolic blood pressure than control mice (P<0.05). The relaxation of aortas to acetylcholine, but not to sodium nitroprusside, was significantly decreased in obese mice and was corrected by both SOD and sodium salicylate (P<0.05). The protein expression of endothelial nitric oxide synthase and Cu/Zn-SOD was significantly decreased in aorta from obese mice (P<0.05). Total p65 NF-kappa B subunit protein expression was not affected by obesity, but the protein expression of NF-kappa B inhibitor I kappa B-alpha was lower in aorta from obese mice (P<0.05). There were no significant differences in the interleukin (IL)-1 beta and IL-6 protein expression between groups. In contrast, the expression of TNF-alpha was significantly increased in aortas from obese mice. Conclusion Our resultssuggest that the reducedantioxidant defense and the local NF-kappa B pathway play an important role in the impairment of endothelium-dependent relaxation in aorta from obese mice. J Hypertens 28: 2111-2119 (C) 2010 Wolters Kluwer Health vertical bar Lippincott Williams & Wilkins.

Effect of chronic supplementation with shark liver oil on immune responses of exercise-trained rats

Previous studies have reported that chronic supplementation with shark liver oil (SLO) improves immune response of lymphocyte, macrophage and neutrophil in animal models and humans. In a similar manner, exercise training also stimulates the immune system. However, we are not aware of any study about the association of exercise and SLO supplementation on immune response. Thus, our main goal was to investigate the effect of chronic supplementation with SLO on immune responses of exercise-trained rats. Male Wistar rats were divided into four groups: sedentary with no supplementation (SED, n = 20), sedentary with SLO supplementation (SEDslo, n = 20), exercised (EX, n = 17) and exercised supplemented with SLO (EXslo, n = 19). Rats swam for 6 weeks, 1.5 h/day, in water at 32 +/- A 1A degrees C, with a load of 6.0% body weight attached to the thorax of rat. Animals were killed 48 h after the last exercise session. SLO supplementation did not change phagocytosis, lysosomal volume, superoxide anion and hydrogen peroxide production by peritoneal macrophages and blood neutrophils. Thymus and spleen lymphocyte proliferation were significantly higher in SEDslo, EX, and EXslo groups compared with SED group (P < 0.05). Gut-associated lymphocyte proliferation, on the other hand, was similar between the four experimental groups. Our findings show that SLO and EX indeed are able to increase lymphocyte proliferation, but their association did not induce further stimulation in the adaptive immune response and also did not modify innate immunity.