Zinc-regulating proteins, ZnT-1, and metallothionein I/II are present in different cell populations in the mouse testis.

Zinc ions play an important role in testis development and spermatogenesis. Thus, nutritional zinc deficiency leads to aberrant testicular development, reduced spermatogenesis, and male sterility. The precise actions of zinc in mediating these functions and the mechanisms by which zinc is itself regulated in the testis, however, have not been adequately elucidated. We have assessed the distribution of the zinc-regulating proteins ZnT-1 and metallothionein I/II (MT I/II) in the mouse seminiferous tubule. Co-labeling for ZnT-1 and MT I/II demonstrated unique patterns of distribution for these proteins, with ZnT-1 present in Sertoli cells in addition to luminal spermatozoa and MT I/II restricted to spermatocytes. These findings were confirmed by dual-label immunofluorescence for ZnT-1 and the Sertoli cell marker, vimentin, and by immunoelectron microscopy. The differential expression patterns of ZnT-1 and MTs support the hypothesis that ZnT-1 and MTs play different roles in the regulation of intracellular zinc in this organ. The specific expression of ZnT-1 in the Sertoli cells, moreover, is consistent with their role in maintaining a nurturing, closely regulated environment for spermatogenesis.     

A lattice relaxation algorithm for three-dimensional Poisson-Nernst-Planck theory with application to ion transport through the gramicidin A channel.

A lattice relaxation algorithm is developed to solve the Poisson-Nernst-Planck (PNP) equations for ion transport through arbitrary three-dimensional volumes. Calculations of systems characterized by simple parallel plate and cylindrical pore geometries are presented in order to calibrate the accuracy of the method. A study of ion transport through gramicidin A dimer is carried out within this PNP framework. Good agreement with experimental measurements is obtained. Strengths and weaknesses of the PNP approach are discussed.     

Characterization of Porins Isolated from the Outer Membrane of Serratia liquefaciens

A major outer membrane protein with an apparent molecular weight of 42 kDa was purified from Serratia liquefaciens grown on Brain Heart Infusion medium. The same protein was obtained when the cells were grown on a synthetic medium supplemented with 2% glucose. The amino acid composition of this protein revealed it to be hydrophilic. The pore-forming ability of the 42-kDa protein was determined by the liposome swelling assay. This assay demonstrated that the protein forms nonspecific channels with a diameter between 1.16 and 1.6 nm. An additional protein with a molecular weight of 47 kDa was obtained on synthetic medium supplemented with maltose. This protein exhibited specific pore-forming ability to maltose and maltodextrins, but was also permeable to other compounds, according to their size. When bacteria were grown on Nutrient Broth medium, two outer membrane proteins with molecular weights of 41 kDa and 42 kDa were produced by the bacteria. All three types of proteins represent monomers of respective oligomers. The monomers did not exhibit pore-forming ability when incorporated into liposomes. We, therefore, propose that the oligomer is the functional unit of a porin capable of forming permeability channels in the outer membrane of Serratia liquefaciens. These results indicate that S. liquefaciens contains several porins exhibiting specific osmoregulation or that are induced by a specific nutrient, where the 42-kDa outer membrane protein of this bacterium is certainly a major porin. Received: 6 July 1998 / Accepted: 19 August 1998     

Renal cell cancer in Israel: Sex and ethnic differences in incidence and mortality, 1980–2004

Background: The causes of renal cell cancer (RCC) remain largely unexplained. While the incidence is generally higher in men than in women, little has been reported on ethnic differences. We examine trends in RCC incidence and mortality rates among Israeli Arab and Jewish populations and compared with the rates in other countries. Methods: Age-adjusted RCC incidence and mortality rates in Israel, during 1980–2004, were calculated by sex and population group, using the National Cancer Registry. They were compared with the United States based on the Surveillance Epidemiology and End Results [SEER] program and the IARC database for international comparisons. Results: While RCC incidence rates in Israel are similar to the United States and the European average, the rates are significantly higher among Israeli Jews than Arabs. Men are affected more than women. Incidence rates over the last 24 years have increased among all men and Jewish women, but not among Arab women. Among men, the incidence rate ratio for Jews to Arabs declined from 3.96 in 1980–1982 to 2.34 in 2001–2004, whereas for women there was no change. The mortality rates were higher among Jews than Arab and among men than women. There were no significant change in the mortality rates and rate ratios. Conclusions: Our findings demonstrate marked ethnic differences in RCC in Israel. The lower incidence among Arabs stands in contrast to the higher prevalence of potential risk factors for RCC in this population group. Genetic factors, diet and other lifestyle factors could play protective roles.     

Studying post depositional damage on Acheulian bifaces using 3-D scanning

In this study, we explore post-depositional damage observed on Acheulian bifacial tools by comparing two assemblages: a collection of archaeological handaxes which shows pronounced damage marks associated with high energy water accumulation system, and an experimental assemblage that was rolled and battered in a controlled simulation experiment. Scanning the two assemblages with a precise 3-D optical scanner and subjecting the measured surfaces to the same mathematical analysis enabled the development of quantitative measures assessing and comparing the degree of damage observed on archaeological and experimental tools. The method presented here enables the definition of morphological patterns typically resulting from battering and different from intentional controlled knapping. The most important kinds of damage included the formation of deep, random ‘notch-like’ scars on the lateral edges and substantial degrees of damage to the tip of the tools, but minimal damage to the artifact's butt. Quantifying the degree of damage and its location and morphological characters allows us to present a method by which post depositional damage on archaeological tools can be measured.     

Self-assembly and Self-replication of Short Amphiphilic β-sheet Peptides

Most self-replicating peptide systems are made of α-helix forming sequences. However, it has been postulated that shorter and simpler peptides may also serve as templates for replication when arranged into well-defined structures. We describe here the design and characterization of new peptides that form soluble β-sheet aggregates that serve to significantly accelerate their ligation and self-replication. We then discuss the relevance of these phenomena to early molecular evolution, in light of additional functionality associated with β-sheet assemblies.     

Development and validation of an isoform-independent monoclonal antibody–based ELISA for measurement of lipoprotein(a)

The study aims were to develop a new isoform-independent enzyme-linked immunoassay (ELISA) for the measurement of lipoprotein(a) [Lp(a)], validate its performance characteristics, and demonstrate its accuracy by comparison with the gold-standard ELISA method and an LC-MS/MS candidate reference method, both developed at the University of Washington. The principle of the new assay is the capture of Lp(a) with monoclonal antibody LPA4 primarily directed to an epitope in apolipoprotein(a) KIV₂ and its detection with monoclonal antibody LPA-KIV9 directed to a single antigenic site present on KIV₉. Validation studies were performed following the guidelines of the Clinical Laboratory Improvement Amendments and the College of American Pathologists. The analytical measuring range of the LPA4/LPA-KIV9 ELISA is 0.27–1,402 nmol/L, and the method meets stringent criteria for precision, linearity, spike and recovery, dilutability, comparison of plasma versus serum, and accuracy. Method comparison with both the gold-standard ELISA and the LC-MS/MS method performed in 64 samples with known apolipoprotein(a) isoforms resulted in excellent correlation with both methods (r=0.987 and r=0.976, respectively). Additionally, the variation in apolipoprotein(a) size accounted for only 0.2% and 2.2% of the bias variation, respectively, indicating that the LPA4/LPA-KIV9 ELISA is not affected by apolipoprotein(a) size polymorphism. Peptide mapping and competition experiments demonstrated that the measuring monoclonal antibodies used in the gold-standard ELISA (a-40) and in the newly developed ELISA (LPA-KIV9) are directed to the same epitope, ⁴⁰⁷⁶LETPTVV⁴⁰⁸², on KIV₉. In conclusion, no statistically or clinically significant bias was observed between Lp(a) measurements obtained by the LPA4/LPA-KIV9 ELISA and those obtained by the gold-standard ELISA or LC-MS/MS, and therefore, the methods are considered equivalent.     

Severe hypoxia induces complete antifolate resistance in carcinoma cells due to cell cycle arrest

Antifolates have a crucial role in the treatment of various cancers by inhibiting key enzymes in purine and thymidylate biosynthesis. However, the frequent emergence of inherent and acquired antifolate resistance in solid tumors calls for the development of novel therapeutic strategies to overcome this chemoresistance. The core of solid tumors is highly hypoxic due to poor blood circulation, and this hypoxia is considered to be a major contributor to drug resistance. However, the cytotoxic activity of antifolates under hypoxia is poorly characterized. Here we show that under severe hypoxia, gene expression of ubiquitously expressed key enzymes and transporters in folate metabolism and nucleoside homeostasis is downregulated. We further demonstrate that carcinoma cells become completely refractory, even at sub-millimolar concentrations, to all hydrophilic and lipophilic antifolates tested. Moreover, tumor cells retained sensitivity to the proteasome inhibitor bortezomib and the topoisomerase II inhibitor doxorubicin, which are independent of cell cycle. We provide evidence that this antifolate resistance, associated with repression of folate metabolism, is a result of the inability of antifolates to induce DNA damage under hypoxia, and is attributable to a hypoxia-induced cell cycle arrest, rather than a general anti-apoptotic mechanism. Our findings suggest that solid tumors harboring a hypoxic core of cell cycle-arrested cells may display antifolate resistance while retaining sensitivity to the chemotherapeutics bortezomib and doxorubicin. This study bears important implications for the molecular basis underlying antifolate resistance under hypoxia and its rational overcoming in solid tumors.     

Accumulation of Trehalose and Sucrose in Cyanobacteria Exposed to Matric Water Stress

The drought-resistant cyanobacteria Phormidium autumnale, strain LPP₄, and a Chroococcidiopsis sp. accumulated trehalose, sucrose, and both trehalose and sucrose, respectively, in response to matric water stress. Accumulated sugar concentrations reached values of up to 6.2 μg of trehalose per μg of chlorophyll in P. autumnale, 6.9 μg of sucrose per μg of chlorophyll in LPP₄, and 4.1 μg of sucrose and 3.2 μg of trehalose per μg of chlorophyll in the Chroococcidiopsis sp. The same sugars were accumulated by these cyanobacteria in similar concentrations under osmotic water stress. Cyanobacteria that did not show drought resistance (Plectonema boryanum and Synechococcus strain PCC 7942) did not accumulate significant amounts of sugars when matric water stress was applied.