Polymorphism of INS VNTR is associated with glutamic acid decarboxylase antibodies and postprandial C-peptide in patients with onset of diabetes after 35 years of age

Variability in the number of tandem repeats of the insulin gene (INS VNTR) is probably involved in the genetic regulation of insulin secretion. The aim of this study was to investigate the association of INS VNTR polymorphism with the presence of glutamic acid decarboxylase antibodies (GADA) and C-peptide levels in patients with the onset of diabetes after 35 years of age. We investigated 117 patients, median of age 63 (range 40-83) years, median of diabetes duration 8 (range 1-30) years; 31 GADA-positive and 86 GADA-negative subjects. INS VNTR polymorphism was typed indirectly using - 23HphI (T/A) polymorphism, which is in complete linkage disequilibrium with INS VNTR. The I/I, I/III and III/III genotypes were found in 22 (71 %), 8 (26 %), 1 (3 %) GADA-positive individuals and in 39 (45 %), 35 (41 %), 12 (14 %) GADA-negative individuals, respectively. The Class I allele and the genotype I/I were significantly associated with the presence of GADA (OR=2.72, CI 95 %=1.29-5.73 and OR=2.95, CI 95 %=1.22-7.13). The presence of Class III allele was significantly associated with a higher level of postprandial C-peptide in GADA-positive subjects, even when regarding the duration of diabetes. Our results of INS VNTR polymorphism in patients with the onset of diabetes after 35 years of age confirm the association of Class I INS VNTR with autoimmune diabetes and the protective effect of Class III INS VNTR on the insulin secretion in GADA-positive subjects.     

Environmental change and rain forests on the Sunda shelf of Southeast Asia: drought, fire and the biological cooling of biodiversity hotspots

Environmental change during the Quaternary period has caused changes in the composition and structure of vegetation on the Sunda shelf of Southeast Asia. Climatic conditions drier than the present, particularly during the peak of the last ice age, led to a reduction in the extent of rain forests. Most recently, there has been a close association between drought and the occurrence of major, rain forest fires. Although many rain forest trees show adaptations to periodic drought, this is not the case for frequent or intense fires. Over evolutionary time-scales, major fires may thus have been largely confined to driver vegetation types, such as monsoon and deciduous forests, and only infrequently penetrated rain forest areas. Continental-scale distribution patterns for rain forest species reveal a number of biodiversity hotspots that are consistent for a broad range of taxonomically unrelated taxa. These biodiversity hotspots account for a relatively small part of the total extent of rain forest; they may also represent ecologically relatively stable areas. This paper discusses the location and extent of biodiversity hotspots on the Sunda shelf within the context of past and present environmental change. It finds that whatever the history of biodiversity hotspots, they are increasingly threatened by contemporary environmental change, notably a trend towards increasingly frequent and intense fires. The paper concludes that the trend is likely to continue, without major changes in those activities that degrade and precondition to fire remaining areas of rain forest.     

H－ras在颊粘膜正常、良性、癌前及恶性变上皮中的表达：免疫组织化学定量研究

本文通过建立图象分析方法对免疫组织化学反应结果进行定量,检测观察Ｈ－ｒａｓ在口腔颊粘膜上皮在正常（Ｎ）、慢性炎症（ＩＦ）、癌旁上皮（ＥＡＣ）和鳞癌（ＳＣＣ）的变化过程中的表达并进行分析。结果显示Ｈ－ｒａｓ在ＳＣＣ组中,以中等分化的ＳＣＣ无论是Ｈ－ｒａｓ表达的量还是细胞阳性率都较高。此外,组织学观察显示,Ｈ－ｒａｓ在处于分化末期但尚未角化的正常上皮细胞中有较高的表达。本文结果显示了Ｈ－ｒａｓ的过表达与上皮细胞的会化程度密切相关。本研究还显示,所采用的阳性区域透光值、平均总透光值及阳性反应区域与阴性反应区域比值可靠并有相关性。这进一步说明了用免疫组化定量方法检测Ｈ－ｒａｓ癌基因表达的精确和可靠性。     

Understanding the immutability of restriction enzymes: crystal structure of BglII and its DNA substrate at 1.5 A resolution

Restriction endonucleases are remarkably resilient to alterations in their DNA binding specificity. To understand the basis of this immutability, we have determined the crystal structure of endonuclease BglII bound to its recognition sequence (AGATCT), at 1. 5 A resolution. We compare the structure of BglII to endonuclease BamHI, which recognizes a closely related DNA site (GGATCC). We show that both enzymes share a similar alpha/beta core, but in BglII, the core is augmented by a beta-sandwich domain that encircles the DNA to provide extra specificity. Remarkably, the DNA is contorted differently in the two structures, leading to different protein-DNA contacts for even the common base pairs. Furthermore, the BglII active site contains a glutamine in place of the glutamate at the general base position in BamHI, and only a single metal is found coordinated to the putative nucleophilic water and the phosphate oxygens. This surprising diversity in structures shows that different strategies can be successful in achieving site-specific recognition and catalysis in restriction endonucleases.     

Isolation and characterization of plasma membrane-associated cortical granules from sea urchin eggs

Cortical granules, which are specialized secretory organelles found in ova of many organisms, have been isolated from the eggs of the sea urchins Arbacia punctulata and Strongylocentrtus pupuratus by a simple, rapid procedure. Electron micropscope examination of cortical granules prepared by this procedure reveals that they are tightly attached to large segments of the plasma membrane and its associated vitelline layer. Further evidence that he cortical granules were associated with these cell surface layers was obtained by (125)I-labeling techniques. The cortical granule preparations were found to be rich in proteoesterase, which was purified 32-fold over that detected in a crude homogenate. Similarly, the specific radioactivity of a (125)I-labeled, surface glycoprotein was increased 40-fold. These facts, coupled with electron microscope observations, indicate the isolation procedure yields a preparation in which both the cortical granules and the plasma membrane-vitelline layer are purified to the same extent. Gel electrophoresis of the membrane-associated cortical granule preparation reveals the presence of at least eight polypeptides. The major polypeptide, which is a glycotprotein of apparent mol wt of 100,000, contains most of the radioactivity introduced by (125)I-labeling of the intact eggs. Lysis of the cortical granules is observed under hypotonic conditions, or under isotonic conditions if Ca(2+) ion is present. When lysis is under isotonic conditions is induced by addition of Ca(2+) ion, the electron-dense contents of the granules remain insoluble. In contrast, hypotonic lysis results in release of the contents of the granule in a soluble form. However, in both cases the (125)I-labeled glycoprotein remains insoluble, presumably because it is a component of either the plasma membrane or the vitelline layer. All these findings indicate that, using this purified preparation, it should be possible to carry out in vitro studies to better define some of the initial, surface-related events observed in vivo upon fertilization.     

Optical recording system based on a fiber optic image conduit: assessment of microscopic activation patterns in cardiac tissue.

Optical recording of transmembrane voltage changes with the use of potentiometric dyes has opened the possibility of determining spatial patterns of electrical activity in excitable tissues. To follow such activation patterns on the cellular/subcellular level in heart cell cultures, a recording system was developed that features both high spatial resolution (4-200 microm) and high temporal resolution (uncertainty in the determination of delays between fast rising signals of +/-1 micros). Central to the system is a fiber optic image conduit consisting of 379 individual optical fibers. At one end the fibers are fused to form an input window that matches the size of the field of view of the microscope. At the other end, the fibers are loose, permitting a selectable subset to be connected to 80 discrete photodetectors. This design allows the sensitive area of the imager to be adapted to regions of interest in a given preparation, thus making optimal use of the limited number of detectors. Furthermore, by using a second fiber optic imager, individual photodetectors can be assigned to different optical ports, thus providing the means for fast and simultaneous dual-emission wavelength measurements. This feature permitted the elimination of motion artifacts arising from the myocytes without the use of contraction-suppressing drugs.     

Physiotherapy as an immunoactive therapy? A pilot study

OBJECTIVES: The aim of this study was to confirm the immunoregulatory and anti-inflammatory changes in the immunologic profile after two months of the facilitation physiotherapy in patients with multiple sclerosis; and to determine whether the changes in the immunologic profile correlate with the changes in dehydroepiandrosterone, the brain microstructure and clinical functions. Design & Setting: A group of 12 patients with multiple sclerosis was examined twice: at the beginning and 2 months later after the patients had undergone the facilitation therapy. Standardized tests evaluating chosen clinical functions (balance, righting, equilibrium and protective reactions, tremor, dysdiadochokinesis, dysmetry, fine hand function and walking), immune parameters (parameters of the humoral and cellular immunity), dehydroepiandrosterone and diffusion tensor imaging (the fractional anisotropy, mean diffusivity) were measured. The patients underwent the facilitation physiotherapy in two sessions lasting two hours each week for two months. Results: All clinical and diffusion tensor imaging parameters significantly improved following the therapy. Without the correction for multiple comparisons, there were significant changes in the IgG, IgG1 subclasses, in the numbers of Neutrophils and Lymphocytes, the T cells (CD3+) absolute number, the T cytotoxic subpopulation (CD3+CD8+) absolute number, B cells (CD19+) and the Natural killer cells. In addition, there was a significant correlation between the changes in the clinical functions and the changes in IgG1 (r=0.67), and between the changes in the mean diffusivity and the changes in CD3+CD8+ absolute (r=-0.61). The changes in the immune parameters and the mentioned correlations were not significant in view of the number of comparisons and thus necessitate further validation. No changes in the dehydroepiandrosterone concentration after the therapy were confirmed. Conclusion: The study suggests new possibilities of physiotherapy to influence the psycho-neuro-endocrine-immune response in patients with multiple sclerosis.     

Uncovering the dynamics of cardiac systems using stochastic pacing and frequency domain analyses

Alternans of cardiac action potential duration (APD) is a well-known arrhythmogenic mechanism which results from dynamical instabilities. The propensity to alternans is classically investigated by examining APD restitution and by deriving APD restitution slopes as predictive markers. However, experiments have shown that such markers are not always accurate for the prediction of alternans. Using a mathematical ventricular cell model known to exhibit unstable dynamics of both membrane potential and Ca²⁺ cycling, we demonstrate that an accurate marker can be obtained by pacing at cycle lengths (CLs) varying randomly around a basic CL (BCL) and by evaluating the transfer function between the time series of CLs and APDs using an autoregressive-moving-average (ARMA) model. The first pole of this transfer function corresponds to the eigenvalue (λ_alt) of the dominant eigenmode of the cardiac system, which predicts that alternans occurs when λ_alt≤−1. For different BCLs, control values of λ_alt were obtained using eigenmode analysis and compared to the first pole of the transfer function estimated using ARMA model fitting in simulations of random pacing protocols. In all versions of the cell model, this pole provided an accurate estimation of λ_alt. Furthermore, during slow ramp decreases of BCL or simulated drug application, this approach predicted the onset of alternans by extrapolating the time course of the estimated λ_alt. In conclusion, stochastic pacing and ARMA model identification represents a novel approach to predict alternans without making any assumptions about its ionic mechanisms. It should therefore be applicable experimentally for any type of myocardial cell.