Effects of Caffeoylquinic Acid Derivatives and C-Flavonoid from Lychnophora ericoides on in vitro Inflammatory Mediator Production

In this study we aimed at evaluating the effect of the major polar constituents of the medicinal plant Lychnophora ericoides on the production of inflammatory mediators produced by LPS-stimulated U-937 cells. The 6,8-di-C-beta-glucosylapigenin (vicenin-2) presented no effect on tumor necrosis factor (TNF)-alpha production, but inhibited, in a dose-dependent manner, the production of prostaglandin (PG) E(2) without altering the expression of cyclooxygenase (COX) -2 protein. 3,5-Dicaffeoylquinic acid and 4,5-dicaffeoylquinic acid, at lower concentrations, had small but significant effects on reducing PG E, levels; at higher doses these compounds stimulated PGE(2) and also TNF-alpha production by the cells. All the caffeoylquinic acid derivatives, in a dose-dependent fashion, were able to inhibit monocyte chemoattractant protein-3 synthesis/release, with 4,5-DCQ being the most potent at the highest tested concentration. These results add important information on the effects of plant natural polyphenols, namely vicenin-2 and caffeoylquinic acid derivatives, on the production of inflammatory mediators by cultured cells.

Protective effect of Calendula officinalis extract against UVB-induced oxidative stress in skin: Evaluation of reduced glutathione levels and matrix metalloproteinase secretion

Background and purpose: Calendula officinalis flowers have long been employed time in folk therapy, and more than 35 properties have been attributed to decoctions and tinctures from the flowers. The main uses are as remedies for burns (including sunburns), bruises and cutaneous and internal inflammatory diseases of several origins. The recommended doses are a function both of the type and severity of the condition to be treated and the individual condition of each patient. Therefore, the present study investigated the potential use of Calendula officinalis extract to prevent UV irradiation-induced oxidative stress in skin. Methods: Firstly, the physico-chemical composition of marigold extract(ME) (hydroalcoholic extract)was assessed and the in vitro antioxidant efficacy was determined using different methodologies. Secondly, the cytotoxicity was evaluated in L929 and HepG2 cells with the MTT assay. Finally, the in vivo protective effect of ME against UVB-induced oxidative stress in the skin of hairless mice was evaluated by determining reduced glutathione (GSH) levels and monitoring the secretion/activity of metalloproteinases. Results and conclusions: The polyphenol, flavonoid, rutin and narcissin contents found in ME were 28.6 mg/g, 18.8 mg/g, 1.6 mg/g and 12.2 mg/g, respectively and evaluation of the in vitro antioxidant activity demonstrated a dose-dependent effect of ME against different radicals. Cytoxicity experiments demonstrated that ME was not cytotoxic for L929 and HepG2 cells at concentrations less than or equal to of 15 mg/mL However, concentrations greater than or equal to 30 mg/mL, toxic effects were observed. Finally, oral treatment of hairless mice with 150 and 300 mg/kg of ME maintained GSH levels close to non-irradiated control mice. In addition, this extract affects the activity/secretion of matrix metalloproteinases 2 and 9 (MMP-2 and -9) stimulated by exposure to UVB irradiation. However, additional studies are required to have a complete understanding of the protective effects of ME for skin. (C) 2009 Elsevier Ireland Ltd. All rights reserved.

Neutrophil activation induced by ArtinM: Release of inflammatory mediators and enhancement of effector functions

The D-mannose binding lectin ArtinM from Artocarpus integrifolia, previously known as KM+ and artocarpin. is considered a stimulant of Th1-type immunity, which is able to confer resistance to some intracellular pathogens. In addition, ArtinM induces neutrophil migration by haptotaxis through simultaneous interactions of its carbohydrate recognition domains (CRDs) with glycans expressed on the extracellular matrix and the neutrophil surface. In the present study, we have expanded the characterization of ArtinM as a neutrophil activator. Exposure of neutrophils to ArtinM for 15 min resulted in tyrosine phosphorylation of intracellular proteins, a process that was selectively inhibited by D-mannose or mannotriose. Shortly after stimulation, neutrophils secreted high levels of LTB(4) and underwent shedding of L-selectin from their surface. Exposure to ArtinM enhanced neutrophil functions, such as respiratory burst and zymozan and Listeria monocytogenes phagocytosis. In addition, ArtinM-stimulated neutrophils displayed increased CXCL-8 secretion and TLR2 gene transcription. These results demonstrate that ArtinM is able to induce potent neutrophil activation, a feature that should be strongly considered in the assessment of the lectin capacity to confer resistance against infections. (C) 2009 Elsevier B.V. All rights reserved.

Vascular permeability, neutrophil migration and edematogenic effects induced by the latex of Cryptostegia grandiflora

Inflammatory responses have been described as occurring after exposure to some latex materials. In this study pro-inflammatory activity in the latex of Cryptostegia grandiflora was investigated. The soluble proteins of the latex (CgLP) were isolated from the whole latex and evaluated by in vivo assays. CgLP induced strong inflammatory activity mediated by neutrophil migration, enlarging vascular permeability and increasing myeloperoxidase activity locally in rats. CgLP-induced inflammation was observed in peritonitis, paw edema and air push models. In addition, CgLP caused hyperemia in a healing model. The peritonitis effect was lost when CgLP was previously boiled suggesting the involvement of proinflammatory proteins. Thioglycollate increased the neutrophil migration induced by CgLP, but not by fMLP Mast cell depletion provoked by 40/80 compound did not modify the course of inflammation triggered by CgLP, being similar to fMLP, which suggested that neutrophil migration was induced by direct mechanism mediated by macrophages. Neutrophil migration stimulated by CgLP was strongly inhibited by Dexamethasone and to a lesser extent by Thalidomide, indicating the involvement of cytokines in mediating neutrophil infiltration. Celecoxib and Indomethacin were inhibitory suggesting the involvement of prostaglandins. Cimetidine was effective only in the initial phase of edema. PCA 4248 was ineffective. It is concluded that the latex of C. grandiflora is a potent inflammatory fluid, and also that laticifer proteins may be implicated in this process. (c) 2008 Elsevier Ltd. All rights reserved.

Effect of peritoneal fluid from patients with minimal/mild endometriosis on progesterone release by human granulosa-lutein cells obtained from infertile patients without endometriosis: A pilot study

Objective: To evaluate the effect of peritoneal fluid (PF) from women without and with minimal/mild endometriosis on progesterone (P) release by cultured human granulosa-lutein cells obtained from infertile patients without endometriosis submitted to ovarian hyperstimulation for in vitro fertilization (IVF). Study design: A pilot study was performed. Human granulosa-lutein cells, obtained from 11 infertile patients without endometriosis (tubal or male factors of infertility) submitted to ovarian hyperstimulation for IVF, were cultured without PF (basal production) and with increasing volumes of steroid-extracted PF samples from 11 patients with endometriosis and 11 patients without endometriosis. Progesterone (P) levels in the media after 72 h culture were measured by chemoluminescence assay. The non-parametric Mann-Whitney-test was used for statistical analysis. Results: PF from patients without endometriosis stimulated P release in a dose-dependent manner up to the dose of 100 mu l/ml (10% concentration) when compared with basal production (without adding PF). P release was similar in cultures stimulated with PF from patients with or without endometriosis at 1% (10 mu l/ml) and 5% (50 ml/ml) concentrations. At 10% concentration, there was a non-statistically significant reduction in progesterone release by granulosa cells stimulated with PF from patients with endometriosis. PF from patients with endometriosis significantly reduced P release at 30% concentration (300 mu l/ml). Conclusions: PF stimulates P release by human granulosa-lutein cells in a dose-dependent manner. However, higher concentrations of PF from patients with minimal/mild endometriosis reduce P release, suggesting it contains factors that may compromise ovarian steroidogenesis. (c) 2007 Elsevier Ireland Ltd. All rights reserved.

Glucocorticoid resistance in dialysis patients may impair the kidney allograft outcome

This study examines in vitro steroid sensitivity in chronic renal failure ( CRF) patients and its influence on the allograft outcome. We determined the inhibitory effect of dexamethasone ( DEX) on concanavalin A ( Con-A)-stimulated peripheral blood mononuclear cell ( PBMC) proliferation, and glucocorticoid receptor` ( GR) number of binding sites ( B-max) and affinity ( K-d) in 28 CRF patients and 40 normal healthy controls. Based on K-d values > 95th percentile from controls, patients were divided into two groups: glucocorticoid resistant ( n = 11) and glucocorticoid sensitive ( n = 17). Patients were followed during 18 months post-transplantation observing acute rejection episodes ( ARE), chronic allograft nephropathy ( CAN), allograft failure and death. The DEX concentration that caused 50% inhibition of Con-A-stimulated PBMC proliferation ( IC50) was higher in CRF than in healthy controls ( 2.2 x 10(-5) +/- 1.0 x 10(-5) versus 8.3 x 10(-6) +/- 4.2 x 10(-6) mol/ L, P = 0.02). Values of Kd ( 12.4 +/- 1.8 versus 7.2 +/- 0.9 nM) and Bmax ( 7.7 +/- 1.1 versus 4.1 +/- 0.3 fmol/ mg protein) were higher in CRF patients ( P = 0.02 and P = 0.001, respectively). There were higher incidences of ARE ( P = 0.02) and CAN ( P = 0.002) in the glucocorticoid-resistant group. Univariate and multivariate logistic regression showed that Kd was an independent predictor of ARE ( OR 8.8, P= 0.03) aswell as of CAN ( OR 16.5, P= 0.01). In conclusion, we observed glucocorticoid resistance in a subgroup of CRF patients undergoing dialysis, which led to a higher morbidity due to ARE and CAN in an 18-month follow-up period.

Non-coding transcript in T cells (NTT): Antisense transcript activates PKR and NF-kappa B in human lymphocytes

T cell activation is a complex process involving many steps and the role played by the non-protein-coding RNAs (ncRNAs) in this phenomenon is still unclear. The non-coding T cells transcript (NTT) is differentially expressed during human T cells activation, but its function is unknown. Here, we detected a 426 m NTT transcript by RT-PCR using RNA of human lymphocytes activated with a synthetic peptide of HIV-1. After cloning, the sense and antisense 426 nt NTT transcripts were obtained by in vitro transcription and were sequenced. We found that both transcripts are highly structured and are able to activate PKR. A striking observation was that the antisense 426 nt NTT transcript is significantly more effective in activating PKR than the corresponding sense transcript. The transcription factor NF-kappa B is activated by PKR through phosphorylation and subsequent degradation of its inhibitor I-kappa B beta. We also found that the antisense 426 nt NTT transcript induces more efficiently the degradation Of I-kappa B beta than the sense transcript. Thus, this study suggests that the role played by NTT in the activation of lymphocytes can be mediated by PKR through NF-kappa B activation. However, the physiological significance of the activity of the antisense 426 nt NTT transcript remains unknown. (c) 2007 Elsevier Inc. All rights reserved.

Microscopic Evidences That Bone Marrow Mononuclear Cell Treatment Improves Sciatic Nerve Regeneration After Neurorrhaphy

Cell therapy constitutes a possibility for improving nerve regeneration, increasing the success of nerve repair. We evaluate the use of mononuclear cells in the regeneration of the sciatic nerve after axotomy followed by end-to-end neurorrhaphy. Forty adult male Wistar rats (250300 g) were divided into four groups: (1) sham, (2) neurorrhaphy: the sciatic nerve was sectioned and repaired using epineural sutures, (3) culture medium: after the suture, received an injection of 10 mu L of culture medium into the nerve, and (4) mononuclear cell: after the suture, a concentration of 3 X 10(6) of mononuclear cell was injected in epineurium region. Mononuclear cells were obtained from the bone marrow aspirates and separated by Ficoll-Hypaque method. The histological analyses were performed at the 4th postoperative day. The sciatic functional index, histological, and morphometric analyzes were used to evaluate nerve regeneration at the 6th postoperative week. Six rats were used for immunohistochemical analysis on the 4th postoperative day. In the group 4, on the fourth day, the histological analysis demonstrated a more accelerated degenerative process and an increase of the neurotrophic factors was observed. In the 6th week, all the morphometric results of the group 4 were statistically better compared with groups 2 and 3. There was a statistically significant improvement in the sciatic functional index for group 4 compared with groups 2 and 3. Mononuclear cells stimulated nerve regeneration, most probably by speeding up the Wallerian degeneration process as well as stimulating the synthesis of neurotrophic factors. Microsc. Res. Tech. 74:355-363, 2011. (C) 2010 Wiley-Liss, Inc.

Clonal Complex Chromosome Aberration in Non-Ossifying Fibroma

Cytogenetic information of non-ossifying fibromas (NOFs) is exceptionally limited. This fact relies, in part, on their benign nature but mainly because most cases evolve undetected or there is no need for surgical intervention. We report the case of a NOF arising in the left tibia of a 14-year-old male with an invariable clonal translocation. The karyotype was denoted as 42-46,XY,t(11;3;14)(q23;p21;p11). There are only two previous reported cases of clonally aberrant NOF. Records from additional cases will be essential to assess whether consistent karyotypic aberrations define this lesion. Pediatr Blood Cancer 2010;54:764 767. (C) 2010 Wiley-Liss, Inc.

Effects of six carbohydrate sources on diet digestibility and postprandial glucose and insulin responses in cats

The effects of diets with different starch sources on the total tract apparent digestibility and glucose and insulin responses in cats were investigated. Six experimental diets consisting of 35% starch were extruded, each containing one of the following ingredients: cassava flour, brewers rice, corn, sorghum, peas, or lentils. The experiment was carried out on 36 cats with 6 replications per diet in a completely randomized block design. The brewers rice diet offered greater DM, OM, and GE digestibility than the sorghum, corn, lentil, and pea diets (P < 0.05). For starch digestibility, the brewers rice diet had greater values (98.6%) than the sorghum (93.9%), lentil (95.2%), and pea (96.3%) diets (P < 0.05); however, starch digestibility was > 93% for all the diets, proving that despite the low carbohydrate content of carnivorous diets, cats can efficiently digest this nutrient when it is properly processed into kibble. Mean and maximum glucose concentration and area under the glucose curve were greater for the corn-based diet than the cassava flour, sorghum, lentil, and pea diets (P < 0.05). The corn-based diets led to greater values for the mean glucose incremental concentration (10.2 mg/dL), maximum glucose incremental concentration (24.8 mg/dL), and area under the incremental glucose curve (185.5 mg.dL(-1).h(-1)) than the lentil diet (2.9 mg/dL, 3.1 mg/dL, and -40.4 mg.dL(-1).h(-1), respectively; P < 0.05). When compared with baseline values, only the corn diet stimulated an increase in the glucose response, occurring at 4 and 10 h postmeal (P < 0.05). The corn-based diet resulted in greater values for maximum incremental insulin concentration and area under the incremental insulin curve than the lentil-based diet (P < 0.05). However, plasma insulin concentrations rose in relation to the basal values for cats fed corn, sorghum, pea, and brewers rice diets (P < 0.05). Variations in diet digestibility and postprandial response can be explained by differences in the chemical composition of the starch source, including fiber content and granule structure, and also differences in the chemical compositions of the diets. The data suggest that starch has less of an effect on the cat postprandial glucose and insulin responses than on those of dogs and humans. This can be explained by the metabolic peculiarities of felines, which may slow and prolong starch digestion and absorption, leading to the delayed, less pronounced effects on their blood responses.

Experimental infection of dogs (Canis familiaris) with sporulated oocysts of Neospora caninum

Neospora caninum is widely distributed in the world and this parasite is one of the major causes of abortion in cattle. Dogs and coyotes are definitive hosts of N. caninum and several species of domestic and wild animals are intermediate hosts. Dogs can become infected by the ingestion of tissues containing cysts and then excrete oocysts. It is not yet known whether sporulated oocysts are able to induce a patent infection in dogs, i.e. a shedding of N. caninum oocysts in feces. The objective of this study was to experimentally examine the infection of dogs by sporulated oocysts. The oocysts used in the experiment were obtained by feeding dogs with brain of buffaloes (Bubalus bubalis) positive for anti-N. caninum antibodies by indirect fluorescent antibody test (IFAT >= 200). Oocysts shed by these dogs were confirmed to be N. caninum by molecular methods and by bioassay in gerbils, and sporulated N. caninum oocysts were used for the oral infection of four dogs. The dogs were 8 weeks old and negative for antibodies to N. caninum and Toxoplasma gondii. Dogs 1 and 4 received an inoculum of 10,000 sporulated oocysts each; dog 2 an inoculum of 5000 sporulated oocysts and dog 3 received 1000 sporulated oocysts of N. caninum. The total feces excreted by these dogs were collected and examined daily for a period of 30 days. No oocysts were found in their feces. The dogs were monitored monthly for a 6-month period to observe a possible seroconversion and when this occurred the animals were eliminated from the experiment. Dogs 1 and 4 seroconverted 1 month after the infection with titer, in the IFAT, of 1600 and 800, respectively; the other two dogs presented no seroconvertion during the 6-month period. Dogs 1 and 2 were euthanized 180 days after infection and were examined for the detection of N. caninum in tissues (brain, muscle, lymph node, liver, lung, heart and bone marrow) by immunohistochemistry and PCR with negative results in both techniques. Bioassay in gerbils with brain of these dogs was also performed and again the results were negative. In conclusion, dogs infected with sporulated oocysts of N. caninum were not able to shed oocysts in feces. However, a higher dose of infection stimulated the production of antibodies against N. caninum in the dogs. (C) 2010 Elsevier B.V. All rights reserved.

Anandamide prior to sensitization increases cell-mediated immunity in mice

The endocannabinoid system has become a topic of great interest in pharmacology due to its remarkable distribution in mammal organisms and capacity to play a modulatory role on several physiological systems, including modulation of immunity. Many studies have shown that administration of cannabinoids causes inhibitory effects on immune cells, including decreased proliferation and antigen-presenting cell (APC) costimulatory activity. In contrast, other groups have shown that some cannabinoids might present stimulatory actions on macrophage activity and T cell activation. Therefore, we aimed to investigate whether a treatment in vivo with a low dose of anandamide (0.1 mg/kg) immediately prior to sensitization would have an immunosuppressive or immunostimulatory effect on cell-mediated immunity (Th1 response) in mice. We report here that anandamide, prior to sensitization, was able to increase the Th1 response to ovalbumin in vivo and ex vivo. Anandamide increased delayed type hypersensitivity (DTH), splenocyte proliferation, and IFN-gamma production in a co-culture of adherent and non-adherent splenocytes. Moreover, anandamide prior to sensitization increased both the expression of DC co-stimulatory molecules (CD80/CD86) and IL-12/IL23 (p40) production ex vivo. We have also assessed direct effects of anandamide in the IFN-gamma/IL-4 balance of ConA-stimulated splenocytes in vitro. Anandamide at nanomolar concentrations increased the production of IFN-gamma, while such production decreased at micromolar range. Thus, anandamide induced both the increment of DC activation and IFN-gamma production, which are likely the mechanisms involved in the increase of Th1 response reported here. (C) 2010 Elsevier B.V. All rights reserved.

Dual origin of mesenchymal stem cells contributing to organ growth and repair

In many adult tissues, mesenchymal stem cells (MSCs) are closely associated with perivascular niches and coexpress many markers in common with pericytes. The ability of pericytes to act as MSCs, however, remains controversial. By using genetic lineage tracing, we show that some pericytes differentiate into specialized tooth mesenchyme-derived cells-odontoblasts-during tooth growth and in response to damage in vivo. As the pericyte-derived mesenchymal cell contribution to odontoblast differentiation does not account for all cell differentiation, we identify an additional source of cells with MSC-like properties that are stimulated to migrate toward areas of tissue damage and differentiate into odontoblasts. Thus, although pericytes are capable of acting as a source of MSCs and differentiating into cells of mesenchymal origin, they do so alongside other MSCs of a nonpericyte origin. This study identifies a dual origin of MSCs in a single tissue and suggests that the pericyte contribution to MSC-derived mesenchymal cells in any given tissue is variable and possibly dependent on the extent of the vascularity.

Comparison between computed tomography and clinical evaluation in tumour/node stage and follow-up of oral cavity and oropharyngeal cancer

Objectives: The aim was to verify the concordance of CT evaluation among four radiologists (two oral and maxillofacial and two medical radiologists) at the TN (tumour/node) stage and in the follow-up of oral cavity and oropharyngeal cancer patients. The study also compared differences between clinical and CT examinations in determining the TN stage. Methods: The following clinical and tomographic findings of 15 non-treated oral cavity and oropharyngeal cancer patients were compared: tumour size, bone invasion and lymph node metastases. In another 15 patients, who had previously been treated, a clinical and tomographic analysis comparison for the presence of tumoural recurrence, post-therapeutic changes in muscles and lymph node metastases was performed. The concordances of tomographic evaluation between the radiologists were analysed using the kappa index. Results: Significant agreement was verified between all radiologists for the T stage, but not for the N stage. In the group of treated patients, CT disclosed post-therapeutic changes in muscles, tumour recurrence and lymph node metastases, but no concordance for the detection of lymph node metastases was found between radiologists. In the first group, for all radiologists, no concordance was demonstrated between clinical and tomographic staging. CT was effective for delimitating advanced lesions and for detecting lymph node involvement in N0 stage patients. CT revealed two cases of bone invasion not clinically detected. Conclusions: Interprofessional relationships must be stimulated to improve diagnoses, and to promote a multidisciplinary approach to oral cavity and oropharyngeal cancer. Although CT was important in the diagnosis and follow-up of cancer patients, differences between medical and dental analyses should be acknowledged. Dentomaxillofacial Radiology (2010) 39, 140-148. doi: 10.1259/dmfr/69910245

Substance P regulates the expression of matrix metalloproteinases and tissue inhibitors of metalloproteinase in cultured human gingival fibroblasts

Background and Objective: Substance P may play a role in the pathogenesis of periodontal disease; however, its mechanisms of modulation are not clear. This study evaluated the effect of two concentrations of Substance P on the expression of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in cultured human gingival fibroblasts. Materials and Methods: Fibroblasts were stimulated for 48 h with 10(-4) or 10(-9) m Substance P; untreated fibroblasts served as controls. The expression of MMP-1, -2, -3, -7 and -11 and of TIMP-1 and -2 was evaluated using real-time polymerase chain reaction and western blotting. Resulsts: There was a significant, concentration-dependent stimulatory effect of Substance P on MMP-1, -2, -3 and -7 and TIMP-2 gene expression (p < 0.05), and a probable effect on MMP-11 (p = 0.06). At the higher concentration (10(-4) m Substance P), MMP-1, -2, -3, -7 and -11 and TIMP-2 showed the greatest up-regulation; at the lower concentration (10(-9) (M) Substance P), MMP-1, -3 and -7 and TIMP-2 exhibited diminished up-regulation, with MMP-2 and -11 showing down-regulation (p < 0.05). Expression of TIMP-1 was not affected by Substance P (p > 0.05). Western blotting confirmed that Substance P up-regulated MMP-1, -2, -3 and -11 and TIMP-2. MMP-1, -3 and -11 and TIMP-2 showed greater up-regulation at the higher Substance P concentration and diminished up-regulation at the lower concentration. MMP-2 was up-regulated to a similar degree at both Substance P concentrations. Conclusion: In gingival fibroblast cells, Substance P at the higher concentration (10(-4) m) induced greater up-regulation of MMP-1, -3 and -11 and TIMP-2 expression, but at the lower concentration (10(-9) (M)) induced diminished up-regulation, which may represent a mechanism for modulating periodontal breakdown.