Prostaglandin D2 expression is prognostic in high­grade serous ovarian cancer.

To identify biomarkers that could predict response or lack of response to conventional chemotherapy at the time of diagnosis of high­grade serous ovarian carcinoma (HGSOC), the present study compared large­scale gene expression from patients with short or long disease­free survival times, according to the last cycle of chemotherapy, and validated these findings using reverse transcription­quantitative polymerase chain reaction (RT­qPCR) and conventional immunohistochemical (IHC) analysis. Samples were selected for microarray evaluation, at the time of diagnosis, using the following criteria: Identical debulking primary surgery, International Federation of Gynaecology and Obstetrics staging, histological subtype and grade. These were divided into 2 groups, regarding the outcome after 2 years of follow-up. Prostaglandin D2 synthase 21 kDa (brain) (PTGDS) was found to be expressed at a significantly higher level in the tumours of patients with a short disease­free survival time, and this was validated by RT­qPCR in all samples. Furthermore, the study evaluated PGD2, the protein product of the PTGDS gene, in a large cohort of 114 HGSOC patients using the Ventana Benchmark automated platform, and IHC positivity was correlated with clinicopathological data and outcome. The global gene expression analysis identified 1,149 genes that were differentially expressed in microarray data, according to the patient outcome. Further analysis RT­qPCR validated PTGDS gene expression in the same samples (r=0.945; P<0.001). IHC analysis showed an inverse profile, with positivity for PGD2 strongly associated with an increase in disease­free survival (P=0.009), the absence of relapse (P=0.039) and sensitivity to platinum­based therapy (P=0.016). Multiple Cox regression showed that IHC evaluation of PGD2 was also a prognostic marker associated with relapse (hazard ratio, 0.37; P=0.002). Overall, the results showed that IHC evaluation of PGD2 is an independent marker of good prognosis in HGSOC. This finding contributes to our understanding of the mechanism of tumour regulation and to investigations into biomarkers that predict response to chemotherapy.

Gene and protein expression in the reproductive tract of Brazilian Somalis rams.

Brazilian Somalis is a locally-adapted breed of rams raised in tropical climate and native pastures. The present study was conducted to evaluate gene expression and proteome of the reproductive tract of such rams. Samples were collected from testes, epididymides, seminal vesicles and bulbourethral glands of four rams. Expression of clusterin (CLU), osteopontin (OPN) and prostaglandin D2 synthase (PGDS) genes were evaluated in all samples by real-time PCR. Shotgun proteomic analysis was performed using samples from the head, corpus and cauda epididymides and from all other structures as well. Gene ontology terms and protein interactions were obtained from UniProtKB databases and MetaCore v.6.8 platform. CLU trasncripts were detected in the testes, epididymides, seminal vesicles and bulbourethral glands of the Somalis rams. The initial region and body of the epididymis had the greatest CLU expression. OPN mRNA was localized in all tissues of the ram reproductive tract. PGDS mRNA was detected in the testes and epididymides. Lable-free mass spectrometry allowed the identification of 137 proteins in all samples. Proteins of the epididymis head mainly participate in cellular processes and response to stimulus, participating in catalityc activity and binding. Proteins of epididymis body acted as regulatory proteins and in cellular processes, with binding and catalytic activity. Cauda epididymis molecules were associated with cellular processes and regulation, with binding function and catalytic activity as well. Testis proteins were mainly linked to cell processes and response to stimuli, and had catalytic function. Seminal vesicle proteins were involved in regulation and mainly with binding functions. Most bulbourethral gland proteins participated in cellular processes. The present study is the first to evaluate the proteome and gene expressions in the reproductive tract of Brazilian Somalis rams. Such pieces of information bring significant cointribution for the understanding of the reproductive physiology of locally-adapted livestock.

Root Transcriptome Analysis of Wild Peanut Reveals Candidate Genes for Nematode Resistance.

Wild peanut relatives (Arachis spp.) are genetically diverse and were adapted to a range of environments during the evolution course, constituting an important source of allele diversity for resistance to biotic and abiotic stresses. The wild diploid A. stenosperma harbors high levels of resistance to a variety of pathogens, including the root-knot nematode (RKN) Meloidogyne arenaria, through the onset of the Hypersensitive Response (HR). In order to identify genes and regulators triggering this defense response, a comprehensive root transcriptome analysis during the first stages of this incompatible interaction was conducted using Illumina Hi-Seq. Overall, eight cDNA libraries were produced generating 28.2 GB, which were de novo assembled into 44,132 contigs and 37,882 loci. Differentially expressed genes (DEGs) were identified and clustered according to their expression profile, with the majority being downregulated at 6 DAI, which coincides with the onset of the HR. Amongst these DEGs, 27 were selected for further qRT-PCR validation allowing the identification of nematode-responsive candidate genes that are putatively related to the resistance response. Those candidates are engaged in the salycilic (NBS-LRR, lipocalins, resveratrol synthase) and jasmonic (patatin, allene oxidase cyclase) acids pathways, and also related to hormonal balance (auxin responsive protein, GH3) and cellular plasticity and signaling (tetraspanin, integrin, expansin), with some of them showing contrasting expression behavior between Arachis RKN-resistant and susceptible genotypes. As these candidate genes activate different defensive signaling systems, the genetic (HR) and the induced resistance (IR), their pyramidding in one genotype via molecular breeding or transgenic strategy might contribute to a more durable resistance, thus improving the long-term control of RKN in peanut.

The impact of lipocalin-type-prostaglandin-D-synthase as a predictor of kidney disease in patients with type 2 diabetes.

Hypertension and diabetes are clinical conditions which contribute to the development of chronic kidney disease as well as risk factors for cardiovascular events. In recent years, lipocalin-type-prostaglandin-D-synthase (beta trace protein; BTP) has increasingly been studied as an alternative to creatinine for the evaluation of renal function as well as for being a possible biomarker for cardiovascular disease. It is expected that the levels of BTP in patients with cardiovascular disease are elevated, as is the case with patients with renal dysfunction. The objective of this study is to realize a systematic review of the pertinent literature in respect to BTP as a biomarker of renal dysfunction in diabetic patients. Using the database MEDLINE, a search up to year 2014 was conducted using the follow descriptors: "lipocalin type prostaglandin d synthase" AND "diabetes"; "lipocalin type prostaglandin d synthase" and "diabetic nephropathy"; "beta trace protein" AND "diabetes"; "beta trace protein" AND "diabetic nephropathy". The criteria used for inclusion were the presence of the referring to terms in title or abstract and study conducted in humans. About 17 articles were selected, of which six articles were duplicates, and of which six articles did not investigate any possible relationship between the protein (BTP) and either diabetes or nephropathy. The final result yielded five articles to be analyzed. This review found BTP is not influenced by race, by body mass index nor by patient's sex. BTP can be considered as a reliable early biomarker of renal dysfunction in diabetics. BTP is associated with metabolic syndrome and is also associated with greater cardiovascular risk. Prospective data establishing a correlation between BTP and mortality would have been of great interest, but such articles were not found in this review.

Neutrophil gelatinase-associated lipocalin in kidney transplantation: A review.

Neutrophil gelatinase-associated lipocalin (NGAL) is a protein expressed by kidney tubular cells in response to ischemia, but may also be an early indicator of immunological rejection, calcineurin inhibitor toxicity, obstructive nephropathy, subclinical tubulitis or infection. Although there is currently no evidence to support the routine serial measurement of blood or urinary NGAL to detect subclinical acute tubular injury, NGAL has the potential to provide useful information to those that care for kidney transplant recipients (KTRs). First, high urinary or serum NGAL concentrations shortly after transplantation are a predictor of delayed graft function and are associated with reduced graft function at one year. Secondly, among KTRs with previously stable graft function who then suffer acute graft dysfunction, a high urinary NGAL predicts graft loss at one year. If further refined, diagnostic tests based on NGAL levels may provide future useful clinical tools.

In vivo mechanism study of NGAL in rat renal ischemia-reperfusion injury.

This study aimed to determine the protective effect and mechanism of neutrophil gelatinase-associated lipocalin (NGAL) in rat kidney on ischemia/reperfusion injury (I/R). The rat I/R model was set up by cutting one kidney and clamping the contralateral renal pedicle for 45 min. Male SD rats were randomly divided into sham-operation, I/R and NGAL groups. Hematoxylin-eosin staining was performed to observe the renal pathological changes in the 3 groups; serum creatinine (Scr) and blood urea nitrogen (BUN) determined in blood samples taken from the inferior vena cava 24 h after the reperfusion were measured; TUNEL was used to observe the apoptosis of renal tubular epithelial cells; immunohistochemistry was performed to evaluate the expressions of Bax and activated caspase-3; Western blotting was used to determine the expression changes in apoptotic proteins Fas and Bcl-2. Compared with the I/R group, Scr and BUN of the NGAL group were 63.400 ± 11.908 vs 121.857 ± 17.151 µM and 14.840 ± 2.868 vs 28.557 ± 6.434 mM, respectively. The number of apoptotic tubular epithelial cells was reduced (7.800 ± 1.924 vs 15.400 ± 3.049); the expression of renal tissue Fas mRNA of the NGAL group was decreased (2.34 ± 0.51 vs 6.84 ± 2.34); the expression of the Bax protein was lower (7.440 ± 1.640 vs 15.456 ± 1.955%); the expression of the CC3 protein was decreased (3.171 ± 0.321 vs 7.291 ± 1.059%), while the expression of the Bcl-2 protein increased (6.91 ± 1.64 vs 5.30 ± 1.48), P < 0.05. NGAL had a protective effect towards the renal tubular epithelial cells in I/R, and the effect might have been associated with the reduction in apoptosis and the altered expression of apoptotic proteins, which would thereby reduce tissue damage and protect the kidney.

Clinical significance of neutrophil gelatinase-associated lipocalin (NGAL) in colorectal cancer: a meta-analysis.

Growing evidence has implicated that neutrophil gelatinase-associated lipocalin (NGAL) plays a role in a spectrum of human cancers. Several observational studies from different parts of the world have been devoted to elucidate the clinical relationship between NGAL and colorectal cancer. This meta-analysis aimed to explore the overall accuracy of NGAL detection for the diagnosis and prognosis of colorectal cancer. Based on comprehensive literature screening on Pubmed, Ovid, and CNKI databases, our screening covered all published papers until March 2013. The relevant papers were selected according to some stringent inclusion criteria. Essential data were extracted from the recruited papers and further processed by systematic meta-analysis. The meta-analysis included 5 studies for diagnosis of colorectal cancer. Overall, the pooled sensitivity and specificity of all studies were 73% (95%CI=0.69-0.76) and 89% (95%CI=0.85-0.93). The pooled positive likelihood ratio and negative likelihood ratio were 5.41 (95%CI=3.85- 7.59) and 0.37 (95%CI=0.22-0.62). The pooled diagnostic odds ratios was 18.05 (95%CI=11.77-27.69). The area under the summary receiver operating characteristic curve for the diagnosis of colorectal cancer was 0.87. Meanwhile, 3 studies were included to evaluate the prognostic significance of NGAL overexpression in colorectal cancer patients. The pooled hazard ratio was 2.12 (95%CI=1.35-3.33). High level of NGAL predicted poor disease-free survival. Thus, NGAL is a potential biomarker for the diagnosis and prognosis of colorectal cancer.

Rosiglitazone did not induce acute kidney injury in normocholesterolemic rats despite reduction in glomerular filtration rate.

BACKGROUND/AIMS: Rosiglitazone (RGL) has been used to ameliorate lipids homeostasis and also to treat inflammatory diseases. However, RGL may reduce renal blood flow and glomerular filtration rate (GFR) predisposing to acute kidney injury (AKI). We investigated whether the treatment with RGL induces AKI in normocholesterolemic (NC) and hypercholesterolemic (HC) rats. METHODS: We measured GFR by inulin clearance technique and we quantified urinary neutrophil gelatinase-associated lipocalin (uNGAL) in all groups at baseline and during Ang II-stimulated vasoconstriction. Moreover, we evaluated the presence of renal damaged by histologic examination. RESULTS: At baseline, NC and HC had normal and similar GFR. RGL treatment reduced GFR only in NC+RGL. Unexpectedly, HC+RGL showed high levels of uNGAL although GFR was at normal range. During Ang II-stimulated vasoconstriction, all groups showed reduction in GFR to the same range and we found high levels of uNGAL and high score of renal damage in HC and HC+RGL. CONCLUSION: RGL acts distinctly in normocholesterolemia and in hypercholesterolemia. Reduction in GFR provoked by RGL treatment did not allow the diagnosis of AKI in NC even in the presence of ANG II-stimulated vasoconstriction. However, AKI was diagnosed in HC+RGL at baseline although GFR was within normal range.

Intrauterine growth restriction modifies the normal gene expression in kidney from rabbit fetuses.

The aim of this work was to study the effect of intrauterine growth restriction (IUGR) on fetal kidneys. The IUGR was induced by uteroplacental vessels ligature in a model of pregnant rabbit. We centralized the study in the gene expression of essential proteins for fetal kidney development and kidney protection against hypoxia, osmotic stress, and kidney injury. The gene expression of HIF-1α, NFAT5, IL-1ß, NGAL, and ATM were studied by qRT-PCR and Western blot in kidneys from control and IUGR fetuses. Experimental IUGR fetuses were significantly smaller than the control animals (39 vs. 48 g, p<0.05). The number of glomeruli was decreased in IUGR kidneys, without morphological alterations. IUGR increased the gene expression of HIF-1α, NFAT5, IL-1ß, NGAL, and ATM (p<0.05) in kidneys of fetuses undergoing IUGR, suggesting that fetal blood flow restriction produce alterations in gene expression in fetal kidneys.

Angiotensin-(1-7) attenuates diabetic nephropathy in Zucker diabetic fatty rats.

Angiotensin (ANG)-(1-7) is known to attenuate diabetic nephropathy; however, its role in the modulation of renal inflammation and oxidative stress in type 2 diabetes is poorly understood. Thus in the present study we evaluated the renal effects of a chronic ANG-(1-7) treatment in Zucker diabetic fatty rats (ZDF), an animal model of type 2 diabetes and nephropathy. Sixteen-week-old male ZDF and their respective controls [lean Zucker rats (LZR)] were used for this study. The protocol involved three groups: 1) LZR + saline, 2) ZDF + saline, and 3) ZDF + ANG-(1-7). For 2 wk, animals were implanted with subcutaneous osmotic pumps that delivered either saline or ANG-(1-7) (100 ng·kg(-1)·min(-1)) (n = 4). Renal fibrosis and tissue parameters of oxidative stress were determined. Also, renal levels of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), ED-1, hypoxia-inducible factor-1α (HIF-1α), and neutrophil gelatinase-associated lipocalin (NGAL) were determined by immunohistochemistry and immunoblotting. ANG-(1-7) induced a reduction in triglyceridemia, proteinuria, and systolic blood pressure (SBP) together with a restoration of creatinine clearance in ZDF. Additionally, ANG-(1-7) reduced renal fibrosis, decreased thiobarbituric acid-reactive substances, and restored the activity of both renal superoxide dismutase and catalase in ZDF. This attenuation of renal oxidative stress proceeded with decreased renal immunostaining of IL-6, TNF-α, ED-1, HIF-1α, and NGAL to values similar to those displayed by LZR. Angiotensin-converting enzyme type 2 (ACE2) and ANG II levels remained unchanged after treatment with ANG-(1-7). Chronic ANG-(1-7) treatment exerts a renoprotective effect in ZDF associated with a reduction of SBP, oxidative stress, and inflammatory markers. Thus ANG-(1-7) emerges as a novel target for treatment of diabetic nephropathy.

Oviductal fluid proteins associated with the bovine zona pellucida and the effect on in vitro sperm-egg binding, fertilization and embryo development.

Studies have demonstrated that oviductal fluid (ODF) proteins associate with eggs of numerous species including the bovine. In this study, the association of three ODF proteins, the bovine oestrus-associated protein, osteopontin (OPN), lipocalin-type prostaglandin D synthase (L-PGDS), with the bovine zona pellucida (ZP) was demonstrated by immunohistochemistry and western blot. The biological function of ODF derived egg-associated OPN and L-PGDS in sperm binding, fertilization and embryonic development was also explored. In vitro matured bovine oocytes were pre-incubated with ODF collected by cannula from cows in oestrus, or ODF with antibodies to OPN, L-PGDS and bovine serum albumin (BSA). Following incubation, oocytes were inseminated with 1 x 10(5) frozen-thawed spermatozoa, and they were evaluated for sperm binding, fertilization and embryonic development in vitro. Pre-treatment of ODF with antibodies to all of proteins reduced sperm binding to the ZP and fertilization in vitro. Cleavage rates were not significantly different among incubations, but rates of embryo development were significantly decreased. We conclude that antibodies to OPN, L-PGDS and BSA react with oocytes incubated with ODF and inhibit sperm binding, fertilization and embryonic development in vitro, suggesting a potential role of these proteins in these events.

An insight into the sialome of the blood-sucking bug Triatoma infestans, a vector of Chagas' disease.

Triatoma infestans is a hemiptera, vector of Chagas' disease that feeds exclusively on vertebrate blood in all life stages. Hematophagous insects' salivary glands (SG) produce potent pharmacological compounds that counteract host hemostasis, including anticlotting, antiplatelet, and vasodilatory molecules. To obtain a further insight into the salivary biochemical and pharmacological complexity of this insect, a cDNA library from its SG was randomly sequenced. Also, salivary proteins were submitted to two-dimensional gel (2D-gel) electrophoresis followed by MS analysis. We present the analysis of a set of 1534 (SG) cDNA sequences, 645 of which coded for proteins of a putative secretory nature. Most salivary proteins described as lipocalins matched peptide sequences obtained from proteomic results.