Inhibition of Human Neutrophil Elastase by Pentacyclic Triterpenes

Scope

Inhibiting human neutrophil elastase (HNE) is a promising strategy for treating inflammatory lung diseases, such as H1N1 and SARS virus infections. The use of sivelestat, the only clinically registered synthesized HNE inhibitor, is largely limited by its risk of organ toxicity because it irreversibly inhibits HNE. Therefore, potent reversible HNE inhibitors are promising alternatives to sivelestat.

Methods and Results

An in vitro HNE inhibition assay was employed to screen a series of triterpenes. Six pentacyclic triterpenes, but not tetracyclic triterpenes, significantly inhibited HNE. Of these pentacyclic triterpenes, ursolic acid exhibited the highest inhibitory potency (IC₅₀ = 5.51 µM). The HNE inhibitory activity of ursolic acid was further verified using a mouse model of acute smoke-induced lung inflammation. The results of nuclear magnetic resonance and HNE inhibition kinetic analysis showed that the pentacyclic triterpenes competitively and reversibly inhibited HNE. Molecular docking experiments indicated that the molecular scaffold, 28-COOH, and a double bond at an appropriate location in the pentacyclic triterpenes are important for their inhibitory activity.

Conclusion

Our results provide insights into the effects of pentacyclic triterpenes on lung inflammatory actions through reversible inhibition of HNE activity.     

Predictors of Attrition and Immunological Failure in HIV-1 Patients on Highly Active Antiretroviral Therapy from Different Healthcare Settings in Mozambique

In Mozambique, the evaluation of retention in HIV care and ART programmes is limited. To assess rate and predictors of attrition (no retention in care) and HAART effectiveness in HIV-1 infected patients who pay for medication and laboratory testing in Mozambique, we conducted a multicenter survey of HIV-1-infected patients who started HAART during 2002–2006. Cox proportional hazard models were used to assess risk of attrition and of therapy failure. Overall, 142 patients from 16 healthcare centers located in the capital city Maputo were followed-up for 22.2 months (12.1–46.7). The retention rate was 75%, 48% and 37% after one, two and three years, respectively. Risk of attrition was lower in patients with higher baseline CD4 count (P = 0.022) and attending healthcare center 1 (HCC1) (P = 0.013). The proportion of individuals with CD4 count ≤200 cells/µL was 55% (78/142) at baseline and decreased to 6% (3/52) at 36 months. Among the patients with available VL, 86% (64/74) achieved undetectable VL levels. The rate of immunologic failure was 17.2% (95% CI: 12.6–22.9) per 100 person-years. Risk of failure was associated to higher baseline CD4 count (P = 0.002), likely reflecting low adherence levels, and decreased with baseline VL ≥10,000 copies/mL (P = 0.033). These results suggest that HAART can be effective in HIV-1 infected patients from Mozambique that pay for their medication and laboratory testing. Further studies are required to identify the causes for low retention rates in patients with low CD4 counts and to better understand the association between healthcare setting and attrition rate.     

Relative Effects of Road Risk,Habitat Suitability,and Connectivity on Wildlife Roadkills: The Case of Tawny Owls (Strix aluco)

Background

Despite its importance for reducing wildlife-vehicle collisions, there is still incomplete understanding of factors responsible for high road mortality. In particular, few empirical studies examined the idea that spatial variation in roadkills is influenced by a complex interplay between road-related factors, and species-specific habitat quality and landscape connectivity.

Methodology/Principal Findings

In this study we addressed this issue, using a 7-year dataset of tawny owl (Strix aluco) roadkills recorded along 37 km of road in southern Portugal. We used a multi-species roadkill index as a surrogate of intrinsic road risk, and we used a Maxent distribution model to estimate habitat suitability. Landscape connectivity was estimated from least-cost paths between tawny owl territories, using habitat suitability as a resistance surface. We defined 10 alternative scenarios to compute connectivity, based on variation in potential movement patterns according to territory quality and dispersal distance thresholds. Hierarchical partitioning of a regression model indicated that independent variation in tawny owl roadkills was explained primarily by the roadkill index (70.5%) and, to a much lesser extent, by landscape connectivity (26.2%), while habitat suitability had minor effects (3.3%). Analysis of connectivity scenarios suggested that owl roadkills were primarily related to short range movements (<5 km) between high quality territories. Tawny owl roadkills were spatially autocorrelated, but the introduction of spatial filters in the regression model did not change the type and relative contribution of environmental variables.

Conclusions

Overall, results suggest that road-related factors may have a dominant influence on roadkill patterns, particularly in areas like ours where habitat quality and landscape connectivity are globally high for the study species. Nevertheless, the study supported the view that functional connectivity should be incorporated whenever possible in roadkill models, as it may greatly increase their power to predict the location of roadkill hotspots.     

Diabetic Patients Could Do As Well as Non-Diabetic Patients without Inflammation on Peritoneal Dialysis

Background

Diabetic patients on peritoneal dialysis (PD) have lower survival and are more likely complicated with inflammation than their non-diabetic counterparts. Here, we explored the interaction effects between diabetes and inflammation on the survival of PD patients.

Methods

Overall, 2,264 incident patients were enrolled from a retrospective cohort study in China. Patients were grouped according to the baseline levels of high-sensitive C-reactive protein (hsCRP, ≤3 mg/L or >3 mg/L) or serum albumin (SA, ≥38 g/L or <38 g/L). Then, several multivariable adjusted stratified Cox regression models were constructed for these groups to explore the predicted role of diabetes on all-cause or cardiovascular death under inflammatory or non-inflammatory conditions.

Results

Diabetics on PD were more likely to have inflammation than non-diabetics on PD, and they presented with elevated hsCRP (52.7% vs. 47.3%, P = 0.03) or decreased SA (77.9% vs. 62.7%, P < 0.001) levels. After stratification by size of center and controlling for confounding factors, diabetes was found to predict all-cause death in patients with hsCRP >3 mg/L or SA <38 g/L but not in patients with hsCRP ≤3 mg/L or SA ≥38 g/L. Similarly, the presence of diabetes was an indication of cardiovascular death in patients with hsCRP >3 mg/L or SA <38 g/L. However, if further adjusted by baseline cardiovascular disease, the predicted role of diabetes on death related to cardiovascular disease in patients with SA <38 g/L disappeared.

Conclusion

Diabetic patients could do as well as non-diabetic patients without inflammation on peritoneal dialysis. Active strategies should be implemented to improve inflammation status in diabetic patients on PD.     

Bayesian Network Analysis Reveals Alterations to Default Mode Network Connectivity in Individuals at Risk for Alzheimer's Disease

Alzheimer''s disease (AD) is associated with abnormal functioning of the default mode network (DMN). Functional connectivity (FC) changes to the DMN have been found in patients with amnestic mild cognitive impairment (aMCI), which is the prodromal stage of AD. However, whether or not aMCI also alters the effective connectivity (EC) of the DMN remains unknown. We employed a combined group independent component analysis (ICA) and Bayesian network (BN) learning approach to resting-state functional MRI (fMRI) data from 17 aMCI patients and 17 controls, in order to establish the EC pattern of DMN, and to evaluate changes occurring in aMCI. BN analysis demonstrated heterogeneous regional convergence degree across DMN regions, which were organized into two closely interacting subsystems. Compared to controls, the aMCI group showed altered directed connectivity weights between DMN regions in the fronto-parietal, temporo-frontal, and temporo-parietal pathways. The aMCI group also exhibited altered regional convergence degree in the right inferior parietal lobule. Moreover, we found EC changes in DMN regions in aMCI were correlated with regional FC levels, and the connectivity metrics were associated with patients'' cognitive performance. This study provides novel sights into our understanding of the functional architecture of the DMN and adds to a growing body of work demonstrating the importance of the DMN as a mechanism of aMCI.     

Imaging Evidence of Diabetic Choroidopathy In Vivo: Angiographic Pathoanatomy and Choroidal-Enhanced Depth Imaging

Purpose

To describe the pathoanatomy of diabetic choroidopathy (DC) in pre-diagnosed diabetic retinopathy (DR) cases and to provide angiographic and optical evidence for DC using indocyanine green angiography (ICGA) and enhanced depth imaging spectral-domain optical coherence tomography (EDI SD-OCT).

Methods

A retrospective analysis of 80 eyes from 40 DR patients was conducted. In Group One, choroidal vascular abnormalities were evaluated by comparing angiographic findings from simultaneous ICGA with those from fundus fluorescein angiography (FFA). In Group Two, EDI SD-OCT was used to evaluate the subfoveal choroidal thickness (SFCT) and define the choroid boundary in order to acquire the bilateral and symmetric choroidal area (CA). Data were then analyzed by Image Pro Plus 6.0.

Results

In Group One, choroidal abnormalities that were evident using ICGA but not FFA included early hypofluorescent spots in 47 eyes (75.81%), late hyperfluorescent spots in 37 eyes (59.68%), and late choroidal non-perfusion regions in 32 eyes (51.61%). In particular, a significant difference between proliferative DR (PDR) in 17 of 23 eyes (73.91%) and non-PDR in 16 of 39 eyes (41.03%) was observed in late choroidal non-perfusion regions. Eighteen of 31 eyes (58.06%) also exhibited “inverted inflow phenomena.” In Group Two, both the SFCT and CA of eyes with diabetic macular edema and serous macular detachment were significantly greater than those in the other eyes. The CA in panretinal photocoagulation (PRP) treated cases was also greater than that in non-PRP treated cases.

Conclusions

Early hypofluorescent spots, late choroidal non-perfusion regions, inverted inflow phenomena, higher SFCT, and larger CA are qualitative and quantitative indexes for DC. Moreover, the late choroidal non-perfusion region is a risk factor for DC with DR. Our study suggests that the supplemental use of ICGA and EDI SD-OCT with FFA is a better choice for DR patients.     

RKIP Inhibition in Cervical Cancer Is Associated with Higher Tumor Aggressive Behavior and Resistance to Cisplatin Therapy

Cervical cancer is one of the most common cancers in women worldwide, being high-risk group the HPV infected, the leading etiological factor. The raf kinase inhibitory protein (RKIP) has been associated with tumor progression and metastasis in several human neoplasms, however its role on cervical cancer is unclear. In the present study, 259 uterine cervix tissues, including cervicitis, cervical intraepithelial lesions and carcinomas, were analyzed for RKIP expression by immunohistochemistry. We found that RKIP expression was significantly decreased during malignant progression, being highly expressed in non-neoplastic tissues (54% of the samples; 73/135), and expressed at low levels in the cervix invasive carcinomas (∼15% (19/124). Following in vitro downregulation of RKIP, we observed a viability and proliferative advantage of RKIP-inhibited cells over time, which was associated with an altered cell cycle distribution and higher colony number in a colony formation assay. An in vitro wound healing assay showed that RKIP abrogation is associated with increased migratory capability. RKIP downregulation was also associated with an increased vascularization of the tumors in vivo using a CAM assay. Furthermore, RKIP inhibition induced cervical cancer cells apoptotic resistance to cisplatin treatment. In conclusion, we described that RKIP protein is significantly depleted during the malignant progression of cervical tumors. Despite the lack of association with patient clinical outcome, we demonstrate, in vitro and in vivo, that loss of RKIP expression can be one of the factors that are behind the aggressiveness, malignant progression and chemotherapy resistance of cervical cancer.     

A Country-Wide Study of Spoligotype and Drug Resistance Characteristics of Mycobacterium tuberculosis Isolates from Children in China

Background

Tuberculosis (TB) is still a big threat to human health, especially in children. However, an isolation of Mycobacterium tuberculosis culture from pediatric cases remains a challenge. In order to provide some scientific basis for children TB control, we investigated the genotyping and drug resistance characteristics of M. tuberculosis isolates from pediatric cases in China.

Methodology/Principal Findings

In this study, a total of 440 strains including 90 from children (<15 years), 159 from adolescents (15–18 years) and 191 from adults (>18 years) isolated in 25 provinces across China were subjected to spoligotyping and drug susceptibility testing. As a result, Beijing family strains were shown to remain predominant in China (85.6%, 81.1% and 75.4% in three above groups, respectively), especially among new children cases (91.0% vs. 69.6% in previously treated cases, P = 0.03). The prevalence of the Beijing genotype isolates was higher in northern and central China in the total collection (85.1% in northern and 83.9% in central vs. 61.6% in southern China, P<0.001) and a similar trend was seen in all three age groups (P = 0.708, <0.001 and 0.025, respectively). In adolescents, the frequencies of isoniazid (INH)-resistant and ethambutol (EMB)-resistant isolates were significantly higher among Beijing strains compared to non-Beijing genotype strains (P = 0.028 for INH and P = 0.027 for EMB). Furthermore, strong association was observed between resistance to rifampicine (RIF), streptomycin (STR) and multidrug resistance (MDR) among Beijing compared to non-Beijing strains in previously treated cases of children (P = 0.01, 0.01 and 0.025, respectively).

Conclusion/Significance

Beijing family was more prevalent in northern and central China compared to southern China and these strains were predominant in all age groups. The genetic diversity of M. tuberculosis isolates from children was similar to that found in adolescents and adults. Beijing genotype was associated with RIF, STR and MDR resistance in previously treated children.     

Genome sequence of the clover-nodulating Rhizobium leguminosarum bv. trifolii strain TA1

Rhizobium leguminosarum bv. trifolii strain TA1 is an aerobic, motile, Gram-negative, non-spore-forming rod that is an effective nitrogen fixing microsymbiont on the perennial clovers originating from Europe and the Mediterranean basin. TA1 however is ineffective with many annual and perennial clovers originating from Africa and America. Here we describe the features of R. leguminosarum bv. trifolii strain TA1, together with genome sequence information and annotation. The 8,618,824 bp high-quality-draft genome is arranged in a 6 scaffold of 32 contigs, contains 8,493 protein-coding genes and 83 RNA-only encoding genes, and is one of 20 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Community Sequencing Program.     

Genome sequence of Ensifer meliloti strain WSM1022; a highly effective microsymbiont of the model legume Medicago truncatula A17

Ensifer meliloti WSM1022 is an aerobic, motile, Gram-negative, non-spore-forming rod that can exist as a soil saprophyte or as a legume microsymbiont of Medicago. WSM1022 was isolated in 1987 from a nodule recovered from the roots of the annual Medicago orbicularis growing on the Cyclades Island of Naxos in Greece. WSM1022 is highly effective at fixing nitrogen with M. truncatula and other annual species such as M. tornata and M. littoralis and is also highly effective with the perennial M. sativa (alfalfa or lucerne). In common with other characterized E. meliloti strains, WSM1022 will nodulate but fixes poorly with M. polymorpha and M. sphaerocarpos and does not nodulate M. murex. Here we describe the features of E. meliloti WSM1022, together with genome sequence information and its annotation. The 6,649,661 bp high-quality-draft genome is arranged into 121 scaffolds of 125 contigs containing 6,323 protein-coding genes and 75 RNA-only encoding genes, and is one of 100 rhizobial genomes sequenced as part of the DOE Joint Genome Institute 2010 Genomic Encyclopedia for Bacteria and Archaea-Root Nodule Bacteria (GEBA-RNB) project.