Hepcidin-25 in Diabetic Chronic Kidney Disease Is Predictive for Mortality and Progression to End Stage Renal Disease

Background

Anemia is common and is associated with impaired clinical outcomes in diabetic chronic kidney disease (CKD). It may be explained by reduced erythropoietin (EPO) synthesis, but recent data suggest that EPO-resistance and diminished iron availability due to inflammation contribute significantly. In this cohort study, we evaluated the impact of hepcidin-25—the key hormone of iron-metabolism—on clinical outcomes in diabetic patients with CKD along with endogenous EPO levels.

Methods

249 diabetic patients with CKD of any stage, excluding end-stage renal disease (ESRD), were enrolled (2003–2005), if they were not on EPO-stimulating agent and iron therapy. Hepcidin-25 levels were measured by radioimmunoassay. The association of hepcidin-25 at baseline with clinical variables was investigated using linear regression models. All-cause mortality and a composite endpoint of CKD progression (ESRD or doubling of serum creatinine) were analyzed by Cox proportional hazards models.

Results

Patients (age 67 yrs, 53% male, GFR 51 ml/min, hemoglobin 131 g/L, EPO 13.5 U/L, hepcidin-25 62.0 ng/ml) were followed for a median time of 4.2 yrs. Forty-nine patients died (19.7%) and forty (16.1%) patients reached the composite endpoint. Elevated hepcidin levels were independently associated with higher ferritin-levels, lower EPO-levels and impaired kidney function (all p<0.05). Hepcidin was related to mortality, along with its interaction with EPO, older age, greater proteinuria and elevated CRP (all p<0.05). Hepcidin was also predictive for progression of CKD, aside from baseline GFR, proteinuria, low albumin- and hemoglobin-levels and a history of CVD (all p<0.05).

Conclusions

We found hepcidin-25 to be associated with EPO and impaired kidney function in diabetic CKD. Elevated hepcidin-25 and EPO-levels were independent predictors of mortality, while hepcidin-25 was also predictive for progression of CKD. Both hepcidin-25 and EPO may represent important prognostic factors of clinical outcome and have the potential to further define “high risk” populations in CKD.     

Unraveling Comparative Anti-Amyloidogenic Behavior of Pyrazinamide and D-Cycloserine: A Mechanistic Biophysical Insight

Amyloid fibril formation by proteins leads to variety of degenerative disorders called amyloidosis. While these disorders are topic of extensive research, effective treatments are still unavailable. Thus in present study, two anti-tuberculosis drugs, i.e., pyrazinamide (PYZ) and D-cycloserine (DCS), also known for treatment for Alzheimer’s dementia, were checked for the anti-aggregation and anti-amyloidogenic ability on Aβ-42 peptide and hen egg white lysozyme. Results demonstrated that both drugs inhibit the heat induced aggregation; however, PYZ was more potent and decelerated the nucleation phase as observed from various spectroscopic and microscopic techniques. Furthermore, pre-formed amyloid fibrils incubated with these drugs also increased the PC12/SH-SY5Y cell viability as compare to the amyloid fibrils alone; however, the increase was more pronounced for PYZ as confirmed by MTT assay. Additionally, molecular docking study suggested that the greater inhibitory potential of PYZ as compare to DCS may be due to strong binding affinity and more occupancy of hydrophobic patches of HEWL, which is known to form the core of the protein fibrils.     

Origin and Evolution of Sulfadoxine Resistant Plasmodium falciparum

The Thailand-Cambodia border is the epicenter for drug-resistant falciparum malaria. Previous studies have shown that chloroquine (CQ) and pyrimethamine resistance originated in this region and eventually spread to other Asian countries and Africa. However, there is a dearth in understanding the origin and evolution of dhps alleles associated with sulfadoxine resistance. The present study was designed to reveal the origin(s) of sulfadoxine resistance in Cambodia and its evolutionary relationship to African and South American dhps alleles. We sequenced 234 Cambodian Plasmodium falciparum isolates for the dhps codons S436A/F, A437G, K540E, A581G and A613S/T implicated in sulfadoxine resistance. We also genotyped 10 microsatellite loci around dhps to determine the genetic backgrounds of various alleles and compared them with the backgrounds of alleles prevalent in Africa and South America. In addition to previously known highly-resistant triple mutant dhps alleles SGEGA and AGEAA (codons 436, 437, 540, 581, 613 are sequentially indicated), a large proportion of the isolates (19.3%) contained a 540N mutation in association with 437G/581G yielding a previously unreported triple mutant allele, SGNGA. Microsatellite data strongly suggest the strength of selection was greater on triple mutant dhps alleles followed by the double and single mutants. We provide evidence for at least three independent origins for the double mutants, one each for the SGKGA, AGKAA and SGEAA alleles. Our data suggest that the triple mutant allele SGEGA and the novel allele SGNGA have common origin on the SGKGA background, whereas the AGEAA triple mutant was derived from AGKAA on multiple, albeit limited, genetic backgrounds. The SGEAA did not share haplotypes with any of the triple mutants. Comparative analysis of the microsatellite haplotypes flanking dhps alleles from Cambodia, Kenya, Cameroon and Venezuela revealed an independent origin of sulfadoxine resistant alleles in each of these regions.     

Unmet need for additional medical care for sick children in mother's view in rural Bangladesh: implications for improving child health services

While a country's health policy aims to provide health services to all who need them, very little in known about unmet need for additional medical care from users' perspectives in Bangladesh. This study examined unmet medical need (defined as whether a mother felt that, to manage sickness, her child had required medical care that was not available, regardless of reasons and medical care sought) of 2123 under-15 sick children by illness and child's socioeconomic characteristics in rural Bangladesh. The 1996 Health and Socioeconomic Survey conducted in Matlab recorded children's chronic (a disease or a condition lasting 3 months or more) and acute (a disease or a condition with a rapid onset and a short, severe course) morbidity, medical care sought to combat illness and unmet needs for additional medical services in mothers' views to manage the illness. The survey also recorded household socioeconomic data. Logistic regression was used to examine the data. The results reveal that unmet needs for additional medical care were 5.4% for children with acute illnesses, and 30.2% for children with chronic illnesses. For chronic illnesses, seeking medical care to manage illness from any health provider outside the home reduced unmet medical needs. Economic inequalities existed for both acute and chronic illnesses: the odds ratio of unmet medical needs for sick children of the least poor households was 0.42 (95% CI: 0.28-0.64) times that for sick children of the very poor households. The critically high unmet needs for children's chronic morbidity reveal that the chronic disease control programme in Bangladesh needs urgent revisiting and strengthening.     

Application of duplex-PCR in rapid and reliable detection of toxigenic Vibrio cholerae in water samples in Thailand

Toxigenic Vibrio cholerae, the cause of cholera, is a native flora of the aquatic environment which is transmitted through drinking water and still remains the leading cause of morbidity and mortality in many developing countries including Thailand. The culture method (CM), which is routinely used for assessing water quality, has not proven as efficient as molecular methods because the notorious pathogen survives in water mostly in a non-culturable state. We employed duplex-polymerase chain reaction (duplex-PCR) for detection of tcpA and ctxA genes in toxigenic V. cholerae, and compared PCR detection with CM in various waters of Khon Kaen Municipality, Thailand. We also evaluated the effect of different pre-PCR conditions on the results of ctxA and tcpA detection including: 1) water filtered and enriched in alkaline peptone water (APW) for 3 h before PCR, 2) water filtered without enrichment before PCR, and 3) use of only enrichment in APW for 6 h before PCR. Of the 96 water samples (taken from waste-water, potable and waste-water from patients' houses, and from rivers) tested, 48 (50%) were positive for ctxA and tcpA by duplex-PCR, whereas only 29 (30%) were positive for V. cholerae by CM. Of the 29 V. cholerae isolated by CM, 2 (7%) were toxigenic V. cholerae belonging to serovar O1, while the rests were non-O1/ non-O139. Results revealed, therefore, that ctxA and tcpA-targeted duplex PCR is more sensitive than CM for detection of toxigenic V. cholerae from water samples because CM detected much less toxigenic V. cholerae than the non-toxigenic V. cholerae. Template DNA as low as 100 fg or 23 cells of V. cholerae in the water sample was detected in duplex PCR. Pre-PCR filtration followed by enrichment for 3 h significantly increase in the efficiency of duplex-PCR detection of toxigenic V. cholerae.     

A conditionally immortalized cell line model for the study of human prostatic epithelial cell differentiation

In the normal human prostate, undifferentiated proliferative cells reside in the basal layer and give rise to luminal secretory cells. There are, however, few epithelial cell lines that have a basal cell phenotype and are able to differentiate. We set out to develop a cell line with these characteristics that would be suitable for the study of the early stages of prostate epithelial cell differentiation. We produced a matched pair of conditionally immortalized prostate epithelial and stromal cell lines derived from the same patient. The growth of these cells is temperature dependent and differentiation can be induced following a rise in culture temperature. Three-dimensional co-cultures of these cell lines elicited gland-like structures reminiscent of prostatic acini. cDNA microarray analysis of the epithelial line demonstrated changes in gene expression consistent with epithelial differentiation. These genes may prove useful as markers for different prostate cell types. The cell lines provide a model system with which to study the process of prostatic epithelial differentiation and stromal-epithelial interactions. This may prove to be useful in the development of differentiation-targeted prostate cancer therapies.     

A kingdom-specific protein domain HMM library for improved annotation of fungal genomes

Background

Mimivirus isolated from A. polyphaga is the largest virus discovered so far. It is unique among all the viruses in having genes related to translation, DNA repair and replication which bear close homology to eukaryotic genes. Nevertheless, only a small fraction of the proteins (33%) encoded in this genome has been assigned a function. Furthermore, a large fraction of the unassigned protein sequences bear no sequence similarity to proteins from other genomes. These sequences are referred to as ORFans. Because of their lack of sequence similarity to other proteins, they can not be assigned putative functions using standard sequence comparison methods. As part of our genome-wide computational efforts aimed at characterizing Mimivirus ORFans, we have applied fold-recognition methods to predict the structure of these ORFans and further functions were derived based on conservation of functionally important residues in sequence-template alignments.

Results

Using fold recognition, we have identified highly confident computational 3D structural assignments for 21 Mimivirus ORFans. In addition, highly confident functional predictions for 6 of these ORFans were derived by analyzing the conservation of functional motifs between the predicted structures and proteins of known function. This analysis allowed us to classify these 6 previously unannotated ORFans into their specific protein families: carboxylesterase/thioesterase, metal-dependent deacetylase, P-loop kinases, 3-methyladenine DNA glycosylase, BTB domain and eukaryotic translation initiation factor eIF4E.

Conclusion

Using stringent fold recognition criteria we have assigned three-dimensional structures for 21 of the ORFans encoded in the Mimivirus genome. Further, based on the 3D models and an analysis of the conservation of functionally important residues and motifs, we were able to derive functional attributes for 6 of the ORFans. Our computational identification of important functional sites in these ORFans can be the basis for a subsequent experimental verification of our predictions. Further computational and experimental studies are required to elucidate the 3D structures and functions of the remaining Mimivirus ORFans.