Human gamma X satellite DNA: an X chromosome specific centromeric DNA sequence

The cosmid clone, CX16-2D12, was previously localized to the centromeric region of the human X chromosome and shown to lack human X-specific satellite DNA. A 1.2 kb EcoRI fragment was subcloned from the CX16-2D12 cosmid and was named 2D12/E2. DNA sequencing revealed that this 1,205 bp fragment consisted of approximately five tandemly repeated DNA monomers of 220 bp. DNA sequence homology between the monomers of 2D12/E2 ranged from 72.8% to 78.6%. Interestingly, DNA sequence analysis of the 2D12/E2 clone displayed a change in monomer unit orientation between nucleotide positions 585–586 from a tail-to-head arrangement to a head-to-tail configuration. This may reflect the existence of at least one inversion within this repetitive DNA array in the centromeric region of the human X chromosome. The DNA consensus sequence derived from a compilation of these 220 bp monomers had approximately 62% DNA sequence similarity to the previously determined 8 satellite DNA consensus sequence. Comparison of the 2D12/E2 and 8 consensus sequences revealed a 20 bp DNA sequence that was well conserved in both DNA consensus sequences. Slot-blot analysis revealed that this repetitive DNA sequence comprises approximately 0.015% of the human genome, similar to that found with 8 satellite DNA. These observations suggest that this satellite DNA clone is derived from a subfamily of satellite DNA and is thus designated X satellite DNA. When genomic DNA from six unrelated males and two unrelated females was cut with SstI or HpaI and separated by pulsed-field gel electrophoresis, no restriction fragment length polymorphisms were observed for either X (2D12/E2) or 8 (50E4) probes. Fluorescence in situ hybridization localized the 2D12/E2 clone to the lateral sides of the primary constriction specifically on the human X chromosome.     

Delayed-type cutaneous hypersensitivity to melanoma antigens and its implication in active specific immunotherapy in cancer

Summary In this study, we explored whether soluble tumor-cell surface-associated antigens (TAA) might be derived from autochthonous as well as allogeneic sources as immunogens for active specific immunotherapy. Using two popular cell membrane-bound antigen extraction techniques (3 M KCl and isotonic-hypotonic NaCl), we examined the immunogenic potential of such TAA and the specificity of immunologic host reactivity through a delayed-type cutaneous hypersensitivity reaction (DTH) as a guideline for their immunogenic potential in a human malignant melanoma model system. We found that either extraction technique could provide soluble TAA from both autochthonous and allogeneic sources capable of eliciting DTH. While evidence of positive DTH with autochthonous TAA reaffirms the immunogenicity of such TAA, the specificity of host reactivity against TAA derived from allogeneic sources is extremely difficult to establish, even with TAA partially purified by column chromatography in Sephadex G-200. Patients exhibited reactivity to other TAA derived from tumors of different histologies and often to more than one component isolated by column chromatography. Furthermore, when a group of melanoma patients was tested against a panel of melanoma antigens in any random combination, DTH to allogeneic TAA was seen in an unpredictable order and with inconsistent frequency. We conclude, therefore, that while autochthonous antigen immunizations may be justified, more careful studies will be necessary to define the antigenic profile of a given tumor (individual specificity vs shared specificity), establish specificity of alloantigens, and devise suitable methods for testing immunologic specificity for alloantigens, before rational immunotherapy with allogeneic tumor antigens will be feasible.     

Protective Effect of Betaine on Respiratory Enzymes

Effects of betaine and NaC1 in various concentrations on the activities of enzymes in tricarboxylic acid cycle (isocitric dehydrogenase, malic dehydrogenase, succinic dehydrogenase and fumarase), terminal oxidation (cytochrome oxidase) and photorespiratory pathway (glycolate oxidase and hydroxypyruvate reductase) have been studied. Betaine, in contrast to electolyte NaC1 was non-inhibitory to these enzymes up to 500 mmol/L. Partial protection against NaC1 inhibition to the activities of these enzymes were afforded by betaine. These results were consistent with the postulated role of betaine in cytoplasmic osmoregulation. These results showed that betaine was a superior compatible solute.     

九种蹄盖蕨科植物配子体的酯酶同工酶分析

本文培养了蹄盖蕨科９个种即中华蹄盖蕨（Ａｔｈｙｒｉｕｍｓｉｎｅｎｓｅ）带岭蹄盖蕨（Ａ．ｄａｌｉｎｇｅｎｓｅ）、多齿蹄盖蕨（Ａ．ｍｕｌｔｉｄｅｎｔａｔｕｍ）（包括青柄和紫柄２种带岭蹄蕨类型）、东北蛾眉蕨（Ｌｕｎａｔｈｙｒｉｕｍｐｙｃｎｏｓｏｒｕｍ）、朝鲜介蕨（Ｄｒｙｏａｔｈｙｒｉｕｍｃｏｒｅａｎｕｍ）、山冷蕨（Ｃｙｓｔｏｐｔｅｒｉｓｓｕｄｅｔｉｃａ）、假冷蕨（Ｐｓｅｕｄｏｃｙｓｔｏｐｔｅｒｉｓｓｐｉｎｕｌｏｓａ）、欧洲羽节蕨（Ｇｙｍｎｏｃａｒｐｉｕｍｄｒｙｏｐｔｅｒｉｓ）和黑鳞短肠蕨（Ａｌａｎｔｏｄｉａｃｒｅｎａｔａ）的配子体为材料,进行酯酶同工酶的分析,以表明它们的种间差异,其中多齿蹄盖蕨的青柄和紫柄两种类型的酶谱存在明显差异,应考虑紫柄为多齿蹄盖蕨的变型。     

A biofilm model to understand the onset of sulfate reduction in denitrifying membrane biofilm reactors

This work presents a multispecies biofilm model that describes the co‐existence of nitrate‐ and sulfate‐reducing bacteria in the H₂‐based membrane biofilm reactor (MBfR). The new model adapts the framework of a biofilm model for simultaneous nitrate and perchlorate removal by considering the unique metabolic and physiological characteristics of autotrophic sulfate‐reducing bacteria that use H₂ as their electron donor. To evaluate the model, the simulated effluent H₂, UAP (substrate‐utilization‐associated products), and BAP (biomass‐associated products) concentrations are compared to experimental results, and the simulated biomass distributions are compared to real‐time quantitative polymerase chain reaction (qPCR) data in the experiments for parameter optimization. Model outputs and experimental results match for all major trends and explain when sulfate reduction does or does not occur in parallel with denitrification. The onset of sulfate reduction occurs only when the nitrate concentration at the fiber's outer surface is low enough so that the growth rate of the denitrifying bacteria is equal to that of the sulfate‐reducing bacteria. An example shows how to use the model to design an MBfR that achieves satisfactory nitrate reduction, but suppresses sulfate reduction. Biotechnol. Bioeng. 2013; 110: 763–772. © 2012 Wiley Periodicals, Inc.     

A thin‐walled polydimethylsiloxane bioreactor for high‐density hepatocyte sandwich culture

In vitro drug testing requires long‐term maintenance of hepatocyte liver specific functions. Hepatocytes cultured at a higher seeding density in a sandwich configuration exhibit an increased level of liver specific functions when compared to low density cultures due to the better cell to cell contacts that promote long term maintenance of polarity and liver specific functions. However, culturing hepatocytes at high seeding densities in a standard 24‐well plate poses problems in terms of the mass transport of nutrients and oxygen to the cells. In view of this drawback, we have developed a polydimethylsiloxane (PDMS) bioreactor that was able to maintain the long‐term liver specific functions of a hepatocyte sandwich culture at a high seeding density. The bioreactor was fabricated with PDMS, an oxygen permeable material, which allowed direct oxygenation and perfusion to take place simultaneously. The mass transport of oxygen and the level of shear stress acting on the cells were analyzed by computational fluid dynamics (CFD). The combination of both direct oxygenation and perfusion has a synergistic effect on the liver specific function of a high density hepatocyte sandwich culture over a period of 9 days. Biotechnol. Bioeng. 2013; 110: 1663–1673. © 2012 Wiley Periodicals, Inc.