Downregulation of Spry2 by miR-21 triggers malignancy in human gliomas.

Gliomas are associated with high mortality because of their exceedingly invasive character. As these tumors acquire their invasiveness from low-grade tumors, it is very important to understand the detailed molecular mechanisms of invasion onset. Recent evidences suggest the significant role of microRNAs in tumor invasion. Thus, we hypothesized that deregulation of microRNAs may be important for the malignant progression of gliomas. We found that the aberrant expression of miR-21 is responsible for glioma invasion by disrupting the negative feedback circuit of Ras/MAPK signaling, which is mediated by Spry2. Upregulation of miR-21 was triggered by tumor microenvironmental factors such as hyaluronan and growth factors in glioma cells lacking functional phosphatase and tensin homolog (PTEN), but not harboring wild-type PTEN. Consistently with these in vitro results, Spry2 protein levels were significantly decreased in 79.7% of invasive WHO grade II-IV human glioma tissues, but not in non-invasive grade I and normal tissues. The Spry2 protein levels were not correlated with their mRNA levels, but inversely correlated with miR-21 levels. Taken together, these results suggest that the post-transcriptional regulation of Spry2 by miR-21 has an essential role on the malignant progression of human gliomas. Thus, Spry2 may be a novel therapeutic target for treating gliomas.

Discharging patients from the nephrology clinic to primary care--will they get appropriate monitoring of renal function?

BACKGROUND: Chronic kidney disease (CKD) guidelines have been produced to allow affected individuals to be identified early and managed more effectively, thereby reducing cardiovascular risk and slowing the progression of CKD. The guidelines allow patients with stable early CKD, who were previously followed in nephrology clinics, to be discharged back to primary care for monitoring of their CKD. AIM: To determine if patients discharged from the nephrology clinic have appropriate monitoring of renal function in primary care according to the UK CKD guidelines, and if patients are being referred back to the clinic appropriately. METHODS: All patients discharged from a weekly satellite unit general nephrology clinic over a 2-year period were identified (n = 160). Clinic letters, the local laboratory system and direct contact with the general practice were used to determine if the timing of tests of renal function were consistent with the UK CKD guidelines. RESULTS: Most subjects (88%) had CKD Stages 1-3 at the time of discharge (i.e. eGFR > 30 ml/min). After exclusion of patients with an incomplete management plan or insufficient time since discharge (n = 50), 85% of eligible patients (n = 110) had at least one measure of eGFR after discharge. In 65% (n = 84) of these patients, measurement occurred within 1 month of the correct timing according to the guidelines. Four patients were re-referred appropriately. There were no other patients who should have been re-referred due to deteriorating renal function. CONCLUSION: Patients with stable early CKD get appropriate monitoring of renal function after discharge from the nephrology clinic to primary care and are also referred back to the renal clinic appropriately.

Biocompatible membranes do not promote graft recovery following cadaveric renal transplantation.

BACKGROUND: Controversy surrounds the role of biocompatible membrane dialyzers in treatment of acute renal failure. Studies that have shown a benefit have involved critically ill patients where renal recovery and patient mortality are influenced by other comorbid disease. The aim of the present work is to clarify this issue in a more homogeneous population of patients with acute renal failure following cadaveric renal transplantation. METHODS: All patients with delayed graft function between January 1996 and February 1998 were randomized to receive either a biocompatible (BCM, polysulfone) membrane or bioincompatible (BICM, cuprophane) membrane for dialysis until onset of graft function. RESULTS: Forty-one patients were randomized, 23 to receive BCM and 18 BICM. Five patients (2 BCM, 3 BICM; p = NS) with primary non-function of graft were excluded from analysis, leaving 36 cases of acute tubular necrosis (ATN). Patient and donor characteristics were similar in both groups. The BCM group had significantly longer periods of dialysis dependency compared to the BICM group (14 vs 10 days; p = 0.03). There was a tendency towards higher serum creatinine levels in the short term in the BCM group (318 vs 164 micromol/l at 1 month (p = 0.1), 190 vs 169 micromol/l at latest visit (p = 0.07)) and a greater number of acute rejection episodes in the BCM group (3.7 vs 1.7 episodes per 100 days of dialysis dependency, p = 0.1). With an intention-to-treat analysis of all 41 patients originally randomized, there was no significant difference in time to graft recovery between the 2 groups (p = 0.18). CONCLUSIONS: In the setting of ARF posttransplantation, we have found no evidence to support the use of biocompatible membranes for dialysis. Rather, our study provides argument against a large benefit for the use of BCM in the recovery of ARF, as suggested by earlier studies.

Genomics analysis of genes expressed in maize endosperm identifies novel seed proteins and clarifies patterns of zein gene expression.

We analyzed cDNA libraries from developing endosperm of the B73 maize inbred line to evaluate the expression of storage protein genes. This study showed that zeins are by far the most highly expressed genes in the endosperm, but we found an inverse relationship between the number of zein genes and the relative amount of specific mRNAs. Although alpha-zeins are encoded by large multigene families, only a few of these genes are transcribed at high or detectable levels. In contrast, relatively small gene families encode the gamma- and delta-zeins, and members of these gene families, especially the gamma-zeins, are highly expressed. Knowledge of expressed storage protein genes allowed the development of DNA and antibody probes that distinguish between closely related gene family members. Using in situ hybridization, we found differences in the temporal and spatial expression of the alpha-, gamma-, and delta-zein gene families, which provides evidence that gamma-zeins are synthesized throughout the endosperm before alpha- and delta-zeins. This observation is consistent with earlier studies that suggested that gamma-zeins play an important role in prolamin protein body assembly. Analysis of endosperm cDNAs also revealed several previously unidentified proteins, including a 50-kD gamma-zein, an 18-kD alpha-globulin, and a legumin-related protein. Immunolocalization of the 50-kD gamma-zein showed this protein to be located at the surface of prolamin-containing protein bodies, similar to other gamma-zeins. The 18-kD alpha-globulin, however, is deposited in novel, vacuole-like organelles that were not described previously in maize endosperm.

Investigating the hows and whys of DNA endoreduplication.

Endoreduplication is a form of nuclear polyploidization that results in multiple, uniform copies of chromosomes. This process is common in plants and animals, especially in tissues with high metabolic activity, and it generally occurs in cells that are terminally differentiated. In plants, endoreduplication is well documented in the endosperm and cotyledons of developing seeds, but it also occurs in many tissues throughout the plant. It is thought that endoreduplication provides a mechanism to increase the level of gene expression, but the function of this process has not been thoroughly investigated. Numerous observations have been made of endoreduplication, or at least extra cycles of S-phase, as a consequence of mutations in genes controlling several aspects of cell cycle regulation. However, until recently there were few studies directed at the molecular mechanisms responsible for this specialized cell cycle. It is suggested that endoreduplication requires nothing more elaborate than a loss of M-phase cyclin-dependent kinase activity and oscillations in the activity of S-phase cyclin-dependent kinase.

Quantitative trait locus mapping of loci influencing elongation factor 1alpha content in maize endosperm.

The nutritional value of maize (Zea mays) seed is most limited by its protein quality because its storage proteins are devoid of the essential amino acid lysine (Lys). The Lys content of the kernel can be significantly increased by the opaque-2 mutation, which reduces zein synthesis and increases accumulation of proteins that contain Lys. Elongation factor 1alpha (eEF1A) is one of these proteins, and its concentration is highly correlated with the Lys content of the endosperm. We investigated the genetic regulation of eEF1A and the basis for its relationship with other Lys-containing proteins by analyzing the progeny of a cross between a high (Oh51Ao2) and a low (Oh545o2) eEF1A maize inbred. We identified 83 simple sequence repeat loci that are polymorphic between these inbreds; the markers are broadly distributed over the genome (1,402 cM) with an average interval of 17 cM. Genotypic analysis of the F(2) progeny revealed two significant quantitative trait loci that account for 25% of the variance for eEF1A content. One of these is on the short arm of chromosome 4 and is linked with a cluster of 22-kD alpha-zein coding sequences; the other quantitative trait locus is on the long arm of chromosome 7. The content of alpha-zein and gamma-zein was measured in pools of high- and low-eEF1A individuals obtained from this cross, and a higher level of alpha-zein was found to cosegregate with high eEF1A content. Allelic variation at the 22-kD alpha-zein locus may contribute to the difference of eEF1A content between Oh51Ao2 and Oh545o2 by increasing the surface area of protein bodies in the endosperm and creating a more extensive network of cytoskeletal proteins.

Cellular basis of potato spindle tuber viroid systemic movement.

Viroids are small, nontranslatable pathogenic RNAs that replicate autonomously and traffic systemically in their host plants. We have used in situ hybridization to analyze the trafficking pattern of Potato spindle tuber viroid (PSTVd) in tomato and Nicotiana benthamiana. When PSTVd was inoculated onto the stem of a plant, it replicated and trafficked to sink, but not source, leaves. PSTVd was absent from shoot apical meristems. In the flowers of infected plants, PSTVd was present in the sepals, but was absent in the petals, stamens, and ovary. The replicative form of PSTVd was detected in the phloem. Our data demonstrate that (i) PSTVd traffics long distance in the phloem and this trafficking is likely sustained by replication of the viroid in the phloem, and (ii) PSTVd trafficking is governed by plant developmental and cellular factors. The dependency of PSTVd and other viroids on cellular mechanisms for RNA trafficking makes them excellent tools to study such mechanisms.

Comparison of two immunization schedules for a Pseudomonas aeruginosa outer membrane proteins vaccine in burn patients.

The aim of the present study was to compare two immunization schedules for a Pseudomonas aeruginosa outer membrane proteins (OMPs) vaccine in burn patients. In a double-blind, randomized and placebo-controlled clinical trial, 95 adult patients with burn injuries in 10% or greater of total body surface area were randomly allocated to either placebo or immunization groups. Three doses of the vaccine (0.5 or 1.0 mg) were administered intramuscularly at either 3- or 7-day intervals. The vaccine was well tolerated, and no severe adverse reactions were observed in any of the vaccinees. After three immunizations, 88 patients were available for evaluation of serum antibody titers. Elevation of OMPs-specific antibody titers in the immunization groups was significantly higher as compared with the placebo group, and the highest antibody response was obtained by immunization with 1.0-mg doses at 3-day intervals. Conventional blood culture, tissue culture of wound biopsy specimens and a nested polymerase chain reaction (PCR) assay of blood specimens were performed to determine the protective efficacy. The results of the nested PCR indicated that the overall detection rate of P. aeruginosa in blood was significantly lower among immunized patients than placebo patients (6.1 vs. 40.0%, P<0.001). Based on these results, we concluded that the P. aeruginosa OMPs vaccine is safe and highly immunogenic in burn patients, especially with 1.0-mg doses at 3-day intervals, and may be effective in conferring protection against P. aeruginosa bacteremia in burn patients.

Early graft function and patient survival following cadaveric renal transplantation.

BACKGROUND: The influence of events that occur early following renal transplantation such as delayed graft function (DGF) and acute rejection on long-term graft survival has been widely reported, but its association with patient survival has received less attention. METHODS: We studied 589 patients who received their first cadaveric transplants between 1984 and 1993, all of whom received cyclosporine-based immunosuppression and who had a median follow-up of seven years. The following factors were identified, and both univariate and multivariate analyses were used to determine their association with long-term patient and graft survival: age, sex, duration of pretransplant dialysis, primary renal disease, immediate graft function (IGF), DGF, primary nonfunction (PNF), acute rejection, and serum creatinine at 3, 6, and 12 months. RESULTS: Patients with PNF had a poorer survival than those with DGF and IGF (P = 0.01), but there was no difference in survival between DGF and IGF (P = 0.54). Good graft function (serum creatinine of less than 200 mumol/liter) at three months was predictive of better long-term patient survival (P = 0.03). Other factors associated with poor patient outcome were older age, diabetes, adult polycystic kidney disease, male gender, and acute rejection. Cardiovascular disease was the most common cause of death (51.8%). Good graft function at three months (P < 0.001) and an absence of rejection episodes (P = 0.01) were associated with better graft survival. CONCLUSION: Patients with poor levels of early graft function (but not DGF) and those with either acute rejection episodes or early graft loss are at an increased risk of early death. These high-risk groups should be targeted for interventional studies in an attempt to improve patient survival.

Synthesis and antimicrobial activity of novel 3-[(aminopyrimidiniumyl)thio]methyl cephalosporins.

A series of novel cephalosporin compounds which have 3-[(aminopyrimidiniumyl)thio]methyl substituents was synthesized. They show high antimicrobial activity against various bacterial species including Pseudomonas aeruginosa. Structure-activity relationships with various thiopyrimidines, thiopyrimidiniums, bicyclic thiotriazolopyrimidiniums, and bicyclic thioimidazolopyrimidiniums as 3'-substituents were also studied; cephalosporins with quarternized pyrimidinium moieties have better antimicrobial activities than neuteral pyrimidine cephalosporins, and stabilization of the positive charge on the pyrimidinium moieties is essential for better activity. According to semiempirical PM3 calculations, amino and alkylthio substituents on the pyrimidinium rings play a major role in charge stabilization and delocalization.