Flavonoid Profile of the Genista tridentata L., a Species Used Traditionally to Treat Inflammatory Processes.

Ethnopharmacological surveys on Portuguese flora reveal that Genista tridentata L. is a shrub used in traditional medicine for the treatment of various inflammation-related health problems, although scientific support of its benefits is still necessary. In order to establish the anti-inflammatory potential of G. tridentata and support its traditional use, ethanolic extracts of three sections of the plant (root, stem, and leaves) were subjected to in vitro evaluation of anti-inflammatory activity using lipopolysaccharide (LPS)-stimulates macrophages as an inflammation model. Simultaneously, we also aimed to establish the extracts' flavonoids profile. The ethanolic extracts, obtained by Soxhlet extraction, profile of the three sections confirmed their richness in flavonoids, being three prenylated flavonoids isolated and characterized in the root, including a new natural compound, the 3-methoxymundulin. The extracts from the three plant sections showed strong antioxidant activity at the cellular level and significantly inhibit the LPS-triggered NO production by downregulating Nos2 gene transcription and consequently iNOS expression. Additionally, root and stem extracts also decreased the LPS-induced transcription of the pro-inflammatory genes Il1b, Il6, and Ptgs2. Thus, the results support the anti-inflammatory properties attributed to G. tridentate preparations. Relevantly, the roots of the shrub, plant part not used, is an unexplored source of compounds with pharmacological and nutraceutical value.

Methodology and reporting of diagnostic accuracy studies of automated perimetry in glaucoma: evaluation using a standardised approach.

PURPOSE: To evaluate methodological and reporting quality of diagnostic accuracy studies of perimetry in glaucoma and to determine whether there had been any improvement since the publication of the Standards for Reporting of Diagnostic Accuracy (STARD) guidelines. METHODS: A systematic review of English language articles published between 1993 and 2013 reporting the diagnostic accuracy of perimetry in glaucoma. Articles were appraised for methodological quality using the 14-item Quality assessment tool for diagnostic accuracy studies (QUADAS) and evaluated for quality of reporting by applying the STARD checklist. RESULTS: Fifty-eight articles were appraised. Overall methodological quality of these studies was moderate with a median number of QUADAS items rated as 'yes' equal to nine (out of a maximum of 14) (IQR 7-10). The studies were often poorly reported; median score of STARD items fully reported was 11 out of 25 (IQR 10-14). A comparison of the studies published in 10-year periods before and after the publication of the STARD checklist in 2003 found quality of reporting had not substantially improved. CONCLUSIONS: Methodological and reporting quality of diagnostic accuracy studies of perimetry is sub-optimal and appears not to have improved substantially following the development of the STARD reporting guidance. This observation is consistent with previous studies in ophthalmology and in other medical specialities.

Hypoxia-related gene expression profile in childhood acute lymphoblastic leukemia: prognostic implications.

A cellular hypoxic condition is a key event in several human cancers, but knowledge about its role in childhood acute lymphoblastic leukemia (ALL) is very limited. In the present study, the gene expression profile of hypoxia-related genes (HIF1A, CA9, VEGF and SCL2A1) was evaluated in bone marrow samples of 113 pediatric patients. HIF1A mRNA up-regulation was significantly associated with higher 5-year event-free survival rates in patients with B-ALL as well as in the overall ALL population in both univariate and multivariate analysis (p = 0.023 and p = 0.041, respectively). In gene expression analysis, low oxygen levels promoted HIF1A activation in a time-dependent manner, in both ALL cell lines. In vitro cytotoxic assays suggested an initial trend toward hypoxia-related resistance in the first 24 h, but evaluation at later time points (48-72 h) clearly showed that there was no relevant difference in drug response when comparing hypoxic and normal oxygen level conditions. Modulation of mRNA expression of several hypoxia-related genes was also observed after hypoxic exposure in a cell specific manner, suggesting that HIF1A mRNA expression could play a different role in specific subtypes of leukemia. Despite the remaining questions regarding hypoxia-mediated mechanisms, these findings could be helpful to provide new insights into the role of hypoxia in childhood ALL.

Modulation of inhibitory systems to enhance motor rehabilitation: insights for the use of noninvasive brain stimulation

Motor impairment following stroke is a leading cause of disability in adults. Despite advances in motor rehabilitation techniques, many adult stroke survivors never approach full functional recovery. Intriguingly, children exhibit better rehabilitation outcomes when compared to adults suffering from comparable brain injuries, yet the reasons for this remain unclear. A common explanation is that neuroplasticity in adults is substantially limited following stroke, thus constraining the brain's ability to reorganize in response to neurological insult. This explanation, however, does not suffice for there is much evidence suggesting that neuroplasticity in adults is not limited following stroke. We hypothesize that diminished functional recovery in adults is in part due to inhibitory neuronal interactions, such as transcallosal inhibition, that serve to optimize motor performance as the brain matures. Following stroke, these inhibitory interactions pose rigid barriers to recovery by inhibiting activity in the affected regions and hindering recruitment of compensatory pathways. In contrast, children exhibit better rehabilitation outcomes in part because they have not fully developed the inhibitory interactions that impede functional recovery in adults. We suggest that noninvasive brain stimulation can be used in the context of motor rehabilitation following stroke to reduce the effects of existing inhibitory connections, effectively returning the brain to a state that is more amenable to rehabilitation. We conclude by discussing further research to explore this hypothesis and its implications

Modulation of inhibitory systems to enhance motor rehabilitation: insights for the use of noninvasive brain stimulation

Motor impairment following stroke is a leading cause of disability in adults. Despite advances in motor rehabilitation techniques, many adult stroke survivors never approach full functional recovery. Intriguingly, children exhibit better rehabilitation outcomes when compared to adults suffering from comparable brain injuries, yet the reasons for this remain unclear. A common explanation is that neuroplasticity in adults is substantially limited following stroke, thus constraining the brain's ability to reorganize in response to neurological insult. This explanation, however, does not suffice for there is much evidence suggesting that neuroplasticity in adults is not limited following stroke. We hypothesize that diminished functional recovery in adults is in part due to inhibitory neuronal interactions, such as transcallosal inhibition, that serve to optimize motor performance as the brain matures. Following stroke, these inhibitory interactions pose rigid barriers to recovery by inhibiting activity in the affected regions and hindering recruitment of compensatory pathways. In contrast, children exhibit better rehabilitation outcomes in part because they have not fully developed the inhibitory interactions that impede functional recovery in adults. We suggest that noninvasive brain stimulation can be used in the context of motor rehabilitation following stroke to reduce the effects of existing inhibitory connections, effectively returning the brain to a state that is more amenable to rehabilitation. We conclude by discussing further research to explore this hypothesis and its implications.(AU)

Retinol binding protein: a short half life determinant of protein non enzymatic glycation in diabetes.

The non enzymatic glycation of circulating and structural proteins is the main biochemical consequence of the chronic hyperglycaemia of diabetes mellitus. Retinol binding protein (RPB) is a 21K plasma globulin with an half life of 12 hr; its non enzymatic glycation may reflect the variation of short term metabolic control (1-4 days). In this study two blood samples were withdrawn at four days interval from 24 non insulin dependent diabetic patients. Glycated RBP was measured by a two-site immunoradiometric assay and its variations correlated with the correspondent changes in the blood glucose level. A significant correlation (r = 0.471; p less than 0.02) was found between the time 4/time 0 ratios of glycated RBP and the time 4/time 0 ratios of blood glucose. These data suggest that measurement of non-enzymatically glycated RBP may be a useful tool to evaluate the short term state of non enzymatic glycation in diabetes.

Formation and ways of detecting advanced glycation end-products in isolated human glomerular basement membrane and human serum albumin nonenzymatically glycated in vitro.

The formation of advanced glycation end-products (AGE) was investigated in samples of isolated human glomerular basement membrane (hGBM) and human serum albumin (hSA) which had been nonenzymatically glycated in vitro. In order to measure AGE, two methods which differ in principle--the standard spectrofluorescence technique and the spectrophotometric diazonium salt reaction--have been used and compared. Samples of finely homogenized hGBM and hSA were incubated for 10 days in buffer containing 500 mmol/L (9 x 10(3) mg%) and 100 mmol/L (1.8 X 10(3) mg%) D-glucose respectively. At the end of the incubation period, the ambient glucose was removed and the samples were incubated for a further 10 days in glucose-free buffer. During this time, loosely bound sugar was released into the buffer; at the end of the incubation, the emission fluorescence at 440 nm (following continuous excitation at 370 nm) and the absorbance at 492 nm of the glycated hGBM and hSA samples were measured and found to be significantly increased by comparison with native samples (1-way ANOVA: p less than 0.05 with both techniques). Comparison of the two techniques used for AGE detection showed a positive linear correlation (Pearson's correlation coefficient r = 0.714; n = 8; p = 0.02). The released glucose probably originates from reversal of the Schiff base (the first and reversible step of the nonenzymatic glycation reaction), whereas fluorescence and photometric findings prove the presence of stable AGE on both hGBM and hSA. It is concluded that AGE can indeed be formed and detected by two different methods in hGBM (and hSA) subjected to nonenzymatic glycation in vitro.(ABSTRACT TRUNCATED AT 250 WORDS)

Nonenzymic glycation of isolated human glomerular basement membrane changes its physicochemical characteristics and binding properties.

The chronic hyperglycemia in diabetes mellitus enhances the nonenzymic glycation of structural proteins possibly increasing the formation of highly reactive advanced glycation end products (AGE). These protein changes might be involved in tissue-damaging mechanisms leading to diabetic complications, including diabetic nephropathy. To simulate these events, an in vitro model, based on isolated human glomerular basement membrane (hGBM), has been developed. In this study we have investigated the extent of AGE formation and the binding changes induced by the nonenzymic glycation of hGBM. An enriched fraction of hGBM was isolated from normal human kidneys and glycated in vitro by incubation with glucose (500 mmol/l) at 37 degrees C for 10 days. The presence of AGE was investigated by two methods - spectrofluorescence and the diazonium salt reaction - both specific for this type of chemical entity. The binding capacity of glycated hGBM was tested by a 10-day incubation with human insulin, albumin, immunoglobulin G and fibrinogen. Higher relative spectrofluorescence values at 440 nm emission (20.0 +/- 2.0 vs. 12.5 +/- 5.0) and higher absorbance values at 492 nm (0.798 +/- 0.063 vs. 0.429 +/- 0.228) indicated the presence of increased levels of AGE in glycated vs. native hGBM. Insulin and the three proteins were bound to hGBM in increased amounts after its glycation (p less than 0.05). The results obtained in this in vitro model confirm that enhanced nonenzymic glycation of hGBM induces the formation of AGE and possibly, through these compounds, alters its physicochemical and binding properties. This reaction might contribute to the mechanisms eventually leading to diabetic nephropathy.

Inhibition of protein non-enzymic glycation induced by Bendazac.

Enhanced non-enzymic glycation of proteins has been suggested to play a role in the pathogenesis of diabetic microangiopathy. Thus pharmacological inhibition of this reaction could be envisaged to delay the development of late diabetic complications. In the present study we have investigated the effect of a new compound, 1-Benzylindazole-3-oxyacetic acid, Bendazac (BDZ) on the in vitro glycation of soluble proteins (albumin and fibrinogen) and isolated glomerular basement membrane (GBM). The data obtained indicate that BDZ is capable of reducing significantly the glycation of albumin and fibrinogen (p less than 0.001). When present in concentrations usually found in patients undergoing therapy (40-80 micrograms/ml), an inhibitory effect on soluble proteins was also observed. Inhibition of glycation of GBM was found only in the presence of the active metabolite (5 hydroxy BDZ) and at high glucose concentrations. These results suggest that BDZ could interfere with protein non-enzymic glycation and its use in patients with diabetes may be then taken into consideration to evaluate the effect on late diabetic complications.