Mean nuclear area of squamous cell carcinomas of the head and neck using image cytometry.

OBJECTIVE: To determine if mean nuclear area (MNA) in squamous cell carcinomas of the head and neck (SCCHN) correlate with the TNM system and histologic grade. STUDY DESIGN: We measured MNA by image cytometry on 74 primary SCCHN. Fify-five had primary surgery, 16 had radiotherapy, and 3 and both as their primary treatment. RESULTS: The mean MNA was 47.85 microm2 (range, 20.5-84.8). Tumor size, nodal status and histologic grade were, respectively: T1 = 13, T2 = 29, T3 = 18, T4 = 14; N0 = 53, N1 = 15, N2 = 5, N3 = 1; 17 = well, 38 = moderate, 19 = poorly differentiated. Spearman rank and Kruskal-Wallis tests for MNA/histologic grade, MNA/tumor size, MNA/nodal status and MNA/site were calculated; only MNA/node was statistically significant (P<.05). CONCLUSION: MNA increases in primary SCCHN as nodal involvement increases. This may reflect that high MNA may be a biologic marker of primary SCCHN with a poorer prognosis.     

Leaf proteome characterization in the context of physiological and morphological changes in response to copper stress in sorghum

Copper (Cu) is an essential micronutrient required for normal growth and development of plants; however, at elevated concentrations in soil, copper is also generally considered to be one of the most toxic metals to plant cells due to its inhibitory effects against many physiological and biochemical processes. In spite of its potential physiological and economical significance, molecular mechanisms under Cu stress has so far been grossly overlooked in sorghum. To explore the molecular alterations that occur in response to copper stress, the present study was performed in ten-day-old Cu-exposed leaves of sorghum seedlings. The growth characteristics were markedly inhibited, and ionic alterations were prominently observed in the leaves when the seedlings were exposed to different concentrations (0, 100, and 150 µM) of CuSO₄. Using two-dimensional gels with silver staining, 643 differentially expressed protein spots (≥1.5-fold) were identified as either significantly increased or reduced in abundance. Of these spots, a total of 24 protein spots (≥1.5-fold) from Cu-exposed sorghum leaves were successfully analyzed by MALDI-TOF-TOF mass spectrometry. Of the 24 differentially expressed proteins from Cu-exposed sorghum leaves, 13 proteins were up-regulated, and 11 proteins were down-regulated. The abundance of most identified protein species, which function in carbohydrate metabolism, stress defense and protein translation, was significantly enhanced, while that of another protein species involved in energy metabolism, photosynthesis and growth and development were severely reduced. The resulting differences in protein expression patterns together with related morpho-physiological processes suggested that these results could help to elucidate plant adaptation to Cu stress and provide insights into the molecular mechanisms of Cu responses in C₄ plants.     

Curcumin Mimics the Neurocognitive and Anti-Inflammatory Effects of Caloric Restriction in a Mouse Model of Midlife Obesity

Dietary curcumin was studied for its potential to decrease adiposity and reverse obesity- associated cognitive impairment in a mouse model of midlife sedentary obesity. We hypothesized that curcumin intake, by decreasing adiposity, would improve cognitive function in a manner comparable to caloric restriction (CR), a weight loss regimen. 15-month-old male C57BL/6 mice were assigned in groups to receive the following dietary regimens for 12 weeks: (i) a base diet (Ain93M) fed ad libitum (AL), (ii) the base diet restricted to 70% of ad libitum (CR) or (iii) the base diet containing curcumin fed AL (1000 mg/kg diet, CURAL). Blood markers of inflammation, interleukin 6 (IL-6) and C-reactive protein (CRP), as well as an indicator of redox stress (GSH: GSSG ratio), were determined at different time points during the treatments, and visceral and subcutaneous adipose tissue were measured upon completion of the experiment. After 8 weeks of dietary treatment, the mice were tested for spatial cognition (Morris water maze) and cognitive flexibility (discriminated active avoidance). The CR group showed significant weight loss and reduced adiposity, whereas CURAL mice had stable weight throughout the experiment, consumed more food than the AL group, with no reduction of adiposity. However, both CR and CURAL groups took fewer trials than AL to reach criterion during the reversal sessions of the active avoidance task, suggesting an improvement in cognitive flexibility. The AL mice had higher levels of CRP compared to CURAL and CR, and GSH as well as the GSH: GSSG ratio were increased during curcumin intake, suggesting a reducing shift in the redox state. The results suggest that, independent of their effects on adiposity; dietary curcumin and caloric restriction have positive effects on frontal cortical functions that could be linked to anti-inflammatory or antioxidant actions.     

Habitat modelling of juvenile Hilsa Tenualosa ilisha (Clupeiformes) in the coastal ecosystem of the northern Bay of Bengal,Bangladesh

Hilsa shad Tenualosa ilisha comprises the largest and most valuable single fishery of Bangladesh. This study was conducted to identify suitable habitats for Hilsa juvenile in the northern Bay of Bengal along the coast of Bangladesh using Geographical Information Systems (GIS) based Multi-Criteria Evaluation (MCE) of water temperature, dissolved oxygen, pH, salinity, turbidity, current, depth and rainfall. Landsat imagery and 8 thematic layers were analyzed with ENVI and GIS capabilities, and developed as a series of GIS models to identify and prioritize the appropriate habitats for Hilsa juvenile. The study covered 29.484 km² water body and revealed that 56% (16.388 km²) is the most suitable, 21% (6.054 km²) is moderately suitable and 24% (7.042 km²) is not suitable which was consistent with field verification. The assemblage of marinebrackish-freshwater ecosystems with favourable ecological parameters and rainfall patterns are very much important for supporting juvenile Hilsa in the study area. The results are useful for habitat-specific sanctuary development as the efficient conservation may lead to higher production of adult Hilsa and increase the economic benefit to the fishers.     

GerO,a Putative Na+/H+-K+ Antiporter,Is Essential for Normal Germination of Spores of the Pathogenic Bacterium Clostridium perfringens

The genome of the pathogen Clostridium perfringens encodes two proteins, GerO and GerQ, homologous to monovalent cation transporters suggested to have roles in the germination of spores of some Bacillus species. GerO and GerQ were able to transport monovalent cations (K⁺ and/or Na⁺) in Escherichia coli, and gerO and gerQ were expressed only in the mother cell compartment during C. perfringens sporulation. C. perfringens spores lacking GerO were defective in germination with a rich medium, KCl, l-asparagine, and a 1:1 chelate of Ca²⁺ and dipicolinic acid (DPA), but not with dodecylamine, and the defect was prior to DPA release in germination. All defects in gerO spores were complemented by ectopic expression of wild-type gerO. Loss of GerQ had much smaller effects on spore germination, and these effects were most evident in spores also lacking GerO. A modeled structure of GerO was similar to that of the E. coli Na⁺/H⁺ antiporter NhaA, and GerO, but not GerQ contained two adjacent Asp residues thought to be important in the function of this group of cation transporters. Replacement of these adjacent Asp residues in GerO with Asn reduced the protein''s ability to complement the germination defect in gerO spores but not the ability to restore cation transport to E. coli cells defective in K⁺ uptake. Together, these data suggest that monovalent cation transporters play some role in C. perfringens spore germination. However, it is not clear whether this role is directly in germination or perhaps in spore formation.Clostridium perfringens is a gram-positive, spore-forming anaerobic pathogen that causes diseases in animals and humans (13). C. perfringens spores are metabolically dormant, are resistant to many environmental insults, and can survive for long periods. Once conditions are favorable, these spores can germinate, outgrow, return to vegetative growth, and then release toxins and cause disease (14).Bacterial spores initiate germination when they sense a variety of compounds termed germinants, which include nutrients, a 1:1 chelate of Ca²⁺ and pyridine-2,6-dicarboxylic acid (dipicolinic acid [DPA]) (Ca-DPA) and cationic surfactants (21, 31). In spores of Bacillus species, nutrient germinants are sensed by specific germinant receptors located in the spore''s inner membrane, each generally encoded by tricistronic operons of the gerA family. In Bacillus megaterium spores, the interaction of nutrient germinants with their cognate receptors leads to an energy independent efflux of ∼80% of the spore''s depot of Na⁺ and K⁺, as well as much H⁺ efflux causing a rise of the spore core''s pH, all within the first 5 min of germination; this efflux is followed by reuptake of K⁺ by an energy-dependent system (33). The spores'' large depot of Ca-DPA is also released shortly after monovalent cation release. The mechanism of release of monovalent cations during spore germination is not known, but monovalent cation antiporters could be involved somehow in this event. Indeed, a member of the CPA-2 monovalent cation-proton antiporter family of membrane transport proteins (27), GrmA, is essential for germination of B. megaterium ATCC 12872 spores (34), since grmA inactivation makes spores unable to release their DPA and complete germination with a variety of germinants. Similarly, in Bacillus cereus ATCC 10876, a GrmA-type homologue, GerN, is essential for spore germination with inosine but not l-alanine (35), and studies with everted vesicles have shown that GerN possesses electrogenic Na⁺/H⁺-K⁺ antiporter activity (32). The GerN homolog, GerT, also plays a minor role in B. cereus spore germination with inosine, as well as a major role in spore outgrowth under some conditions (29). However, in contrast to these latter results, GrmA-like antiporters appear to have no role in the germination of spores of B. megaterium QM B1551 and Bacillus subtilis (3).In C. perfringens, there is no intact tricistronic gerA-like operon, and the only locus that encodes the three proteins (A, B, and C) of a likely germinant receptor is the gerK locus, comprising a bicistronic gerKA-gerKC operon, and a gerKB gene located just upstream of gerKA-gerKC but in the opposite orientation (16). However, GerKA and GerKC appear able to function in spore germination in the absence of GerKB (23). The lack of a classical GerA-type germinant receptor and the fact that C. perfringens spores germinate with K⁺ ions alone (21), raises the possibility that GrmA-like antiporters might also play some role in C. perfringens spore germination. The genome of C. perfringens strain SM101 has two genes encoding putative GrmA-like antiporters (see Fig. S1 in the supplemental material) that we have termed gerO (CPR0227) and gerQ (CPR1038). Orthologs of the gerO and gerQ genes are also present in the genomes of nine additional C. perfringens strains (http://www.ncbi.nlm.nih.gov/genomes/lproks.cgi). In present study we have constructed gerO, gerQ, and gerO gerQ strains of C. perfringens and have examined the roles of GerO and GerQ in spore germination. The results show that GerO is essential for normal germination of C. perfringens spores, whereas GerQ plays at most only a minor role.     

Two-step affinity purification of the hepatitis C virus ribonucleoprotein complex

Positive-strand RNA viruses replicate their RNA genome within a ribonucleoprotein (RNP) complex that is associated with cellular membranes. We used a two-step method of purification to isolate hepatitis C virus (HCV) RNP complexes from human hepatoma cell line Huh7, which stably expresses HCV subgenomic replicons. The procedure involved hybridization of replicon-expressing cellular lysates with oligonucleotides tagged with biotin and digoxigenin at their respective termini complementary to subgenomic replicon RNA followed by avidin-agarose enrichment of the mixture and subsequent immunoprecipitation of biotin-eluted material with anti-digoxigenin antibody. The immunoprecipitates were immunoblotted with antisera against HCV nonstructural (NS) proteins. The analysis revealed the association of all the HCV NS proteins (NS3, NS4a, NS4b, NS5a, and NS5b) that are encoded by the subgenomic replicon RNA. The HCV RNP complex migrated in a native polyacrylamide gel with an approximate molecular mass of 450 kD. The association of these viral proteins in the RNP complex reinforces the widely acknowledged notion that RNA viruses accomplish replication within a membranous RNP complex.