Morphology, anatomy, palynology and nutlet micromorphology of the rediscovered Turkish endemic Salvia ballsiana (Lamiaceae) and their taxonomic implications

Salvia ballsiana (Rech. fil.) Hedge, previously known only from the type gathering, is a local endemic species from Turkey. The species was first collected from Gölbaşı (Malatya) by E. K. Balls in 1935, and was not collected again until 2008, when we found it in Gerger (Adıyaman). The diagnostic morphological characters of this rare endemic species are discussed, including its anatomical, palynological and nutlet micromorphological features. Morphological characteristics of leaves, calyces, corollas and types of stamens are useful for sectional and specific delimitation in Salvia . Anatomical characters such as number of ray rows in roots, distance between vascular bundles in stems, mesophyll structures in leaves, shape of mid-rib and presence/absence of sclerenchymatic tissue in petioles are of taxonomic significance. In addition, size, shape and ornamentation of pollen grains and nutlets are diagnostic.     

Photobiont association and genetic diversity of the optionally lichenized fungus Schizoxylon albescens

The fungus Schizoxylon albescens occurs both as lichen and as saprobe. Lichenized colonies grow on the bark of Populus tremula; saprotrophic morphs grow on dead Populus branches. We wanted to (1) test whether lichenized and saprotrophic S. albescens are genetically distinct, (2) investigate photobiont association and diversity, (3) investigate the interactions between fungi and algae that occur during co-cultivation and (4) test whether Schizoxylon shows algal selectivity during lichenization. Fungal and algal genetic diversity were investigated for three markers. Algae from lichenized thalli were isolated in axenic cultures, and isolate sequence diversity was compared with algae amplified directly from thallus fragments. Co-culture experiments of fungi and algae were performed to study the morphological interaction patterns. Two distinct phylogenetic units are revealed in S. albescens, which are interpreted as phenotypically cryptic species. The algae are related to Coccomyxa and Pseudococcomyxa, and form two distinct sister clades separating samples isolated in cultures from those amplified directly from thallus fragments, indicating that more easily cultured strains of algae are not necessarily major components of the lichens. Schizoxylon albescens interacts with isolated algal strains, similar to fungal-Coccomyxa symbioses in nature. As the system is maintained without difficulty in culture, it can potentially be an easily controlled lichen symbiosis study system under laboratory conditions.     

Inhibition of LDL oxidation by flavonoids in relation to their structure and calculated enthalpy

Twenty flavonoid compounds of five different subclasses were selected, and the relationship of their structure to the inhibition of low-density lipoprotein (LDL) oxidation in vitro was investigated. The most effective inhibitors, by either copper ion or 2,2'-azobis (2-amidino-propane) dihydrochloride (AAPH) induction, were flavonols and/or flavonoids with two adjacent hydroxyl groups at ring B. In the presence of the later catechol group, the contribution of the double bond and the carbonyl group at ring C was negligible. Isoflavonoids were more effective inhibitors than other flavonoid subclasses with similar structure. Substituting ring B with hydroxyl group(s) at 2' position resulted in a significantly higher inhibitory effect than by substituting ring A or ring B at other positions. The type of LDL inducer had no effect in flavonoids with catechol structure. Calculated heat of formation data (deltadeltaH(f)) revealed that the donation of a hydrogen atom from position 3 was the most likely result, followed by that of a hydroxyl from ring B. Position 3 was favored only in the presence of conjugated double bonds between ring A to ring B. This study makes it possible to assign the contribution of different functional groups among the flavonoid subclasses to in vitro inhibition of LDL oxidation.     

Toward visualization of nanomachines in their native cellular environment

The cellular nanocosm is made up of numerous types of macromolecular complexes or biological nanomachines. These form functional modules that are organized into complex subcellular networks. Information on the ultra-structure of these nanomachines has mainly been obtained by analyzing isolated structures, using imaging techniques such as X-ray crystallography, NMR, or single particle electron microscopy (EM). Yet there is a strong need to image biological complexes in a native state and within a cellular environment, in order to gain a better understanding of their functions. Emerging methods in EM are now making this goal reachable. Cryo-electron tomography bypasses the need for conventional fixatives, dehydration and stains, so that a close-to-native environment is retained. As this technique is approaching macromolecular resolution, it is possible to create maps of individual macromolecular complexes. X-ray and NMR data can be ‘docked’ or fitted into the lower resolution particle density maps to create a macromolecular atlas of the cell under normal and pathological conditions. The majority of cells, however, are too thick to be imaged in an intact state and therefore methods such as ‘high pressure freezing’ with ‘freeze-substitution followed by room temperature plastic sectioning’ or ‘cryo-sectioning of unperturbed vitreous fully hydrated samples’ have been introduced for electron tomography. Here, we review methodological considerations for visualizing nanomachines in a close-to-physiological, cellular context. EM is in a renaissance, and further innovations and training in this field should be fully supported. Robert Feulgen Lecture 2009 presented at the 51st symposium of the Society for Histochemistry in Stubai, Austria, October 7–10, 2009.     

Determination of plasma microRNA for early detection of gastric cancer

Gastric cancer is the fourth most prevalent malignancy worldwide and remains the second most common cause of cancer-related death globally. Understanding the molecular structure of gastric carcinogenesis might identify new diagnostic and therapeutic strategies for this disease. Thus, early detection of gastric cancer is a key measure to reduce the mortality and improve the prognosis of gastric cancer. There have recently been several reports that microRNAs (miRNAs) circulate in highly stable, cell-free forms in blood. Because serum and plasma miRNAs are relatively easy to access, circulating miRNAs also have great potential to serve as non-invasive biomarkers. Although a number of miRNAs associated with gastric cancer have been identified, the underlying mechanism of these miRNAs in tumorigenesis and tumor progression remains to be investigated. The purpose of this study is to identify the potential of serum miRNAs as biomarkers for early detection of gastric cancer patients. RNA was isolated using the High Pure miRNA Isolation Kit (Roche) following the manufacturer’s protocol. cDNA and preamplification protocols were obtained from the isolated plasma miRNAs. The BioMark? 96.96 Dynamic Array (Fluidigm Corporation) for real-time qPCR was used to simultaneously quantite the expression of 740 miRNAs. All statistical analyses were performed using the Biogazelle’s qbase PLUS 2.0 software. In this study, among 740 miRNAs that we analyzed only miR-195-5p was significantly (p < 0.05, fold changes = 13, 3) down-regulated in gastric cancer patients compared with control. We demonstrated that miR-195-5p is a novel tumor suppressor miRNA and may contribute to gastric carcinogenesis. The miRNA expression profile described in this study should contribute to future studies on the role of miRNAs in gastric cancer.     

Novel inner membrane retention signals in Pseudomonas aeruginosa lipoproteins

The ultimate membrane localization and function of most of the 185 predicted Pseudomonas aeruginosa PAO1 lipoproteins remain unknown. We constructed a fluorescent lipoprotein, CSFP^OmlA-ChFP, by fusing the signal peptide and the first four amino acids of the P. aeruginosa outer membrane lipoprotein OmlA to the monomeric red fluorescent protein mCherry (ChFP). When cells were plasmolyzed with 0.5 M NaCl, the inner membrane separated from the outer membrane and formed plasmolysis bays. This permits the direct observation of fluorescence in either the outer or inner membrane. CSFP^OmlA-ChFP was shown to localize in the outer membrane by fluorescence microscopy and immunoblotting analysis of inner and outer membrane fractions. The site-directed substitution of the amino acids at positions +2, +3, and +4 in CSFP^OmlA-ChFP was performed to test the effects on lipoprotein localization of a series of amino acid sequences selected from a panel of predicted lipoproteins. We confirmed Asp⁺² and Lys⁺³ Ser⁺⁴ function as inner membrane retention signals and identified four novel inner membrane retention signals: CK⁺² V⁺³ E⁺⁴, CG⁺² G⁺³ G⁺⁴, CG⁺² D⁺³ D⁺⁴, and CQ⁺² G⁺³ S⁺⁴. These inner membrane retention signals are found in 5% of the 185 predicted P. aeruginosa lipoproteins. Full-length chimeras of predicted lipoproteins PA4370 and PA3262 fused to mCherry were shown to reside in the inner membrane and showed a nonuniform or patchy distribution in the membrane. The optical sectioning of cells producing PA4370^CGDD-ChFP and PA3262^CDSQ-ChFP by confocal microscopy improved the resolution and indicated a helix-like localization pattern in the inner membrane. The method described here permits the in situ visualization of lipoprotein localization and should work equally well for other membrane-associated proteins.     

Macroinvertebrates as unreliable indicators of human disturbance in temporary depression wetlands of the south-western Cape, South Africa

Macroinvertebrates have a successful history of use as indicators of human impact in lotic environments. More recently, macroinvertebrate indices have been recommended for use in certain wetland types. Yet some authors do not recommend macroinvertebrates indices of wetland condition in areas with pronounced natural environmental heterogeneity. Our study provides a preliminary assessment of the feasibility of using macroinvertebrates for bioassessment of temporary isolated depression wetlands in the south-western Cape region of South Africa. We expected natural environmental heterogeneity among wetlands to exert a stronger influence on macroinvertebrates than human disturbance factors. Partitioning of the variation in macroinvertebrate assemblage composition that could be attributed to human disturbance factors (within and adjacent to wetlands), environmental variables and spatio-temporal factors indicated that environmental and spatio-temporal factors independent of human disturbances largely determined assemblage composition, whilst human disturbance played a relatively minor role. Linear regressions of taxon richness/diversity measures, individual families and a collation of metrics against measures of habitat transformation around wetlands and scores from a rapid assessment index of human disturbance revealed poor relationships. The univariate and multivariate patterns observed in this study do not lend themselves to the creation of a macroinvertebrate index of human disturbance for temporary wetlands in the region.     

Assessment of DNA strand breakage by the alkaline COMET assay in dialysis patients and the role of Vitamin E supplementation

Although the role of reactive oxygen species (ROS) in chronic renal failure (CRF) is not definitely demonstrated, a consistent number of observations has provided evidence for the presence of oxidative stress in uremic patients undergoing maintenance dialysis. In order to investigate this hypothesis further and to understand the role of antioxidant supplementation, peripheral blood lymphocytes were taken from 36 dialysis patients before and after Vitamin E supplementation in a dosage of 600 mg per day (2x300 mg) for 14 weeks and examined in the alkaline Comet assay for DNA strand breakage. The results were also compared with those of 36 controls with comparable age, sex, and smoking habits, and with no history of renal disease. The DNA breakage observed in the lymphocytes of patients before Vitamin E supplementation was significantly higher than in the controls (P<0.001) but a clear protective effect of Vitamin E supplementation were observed after 14 weeks of therapy.