Severe hypoglycemia and hip fracture in patients with type 2 diabetes: a nationwide population-based cohort study.

Hypoglycemia is a major concern in glycemic control. Using the Taiwan National Health Insurance Research Database, we found that the risk of hip fracture was associated with emergency or hospitalization visits of severe hypoglycemia in patients with type 2 diabetes; greater visits were associated with higher incidence of hip fracture. INTRODUCTION: The objective of the study was to assess the risk of hip fracture among patients with type 2 diabetes mellitus (T2DM) and severe hypoglycemia. METHODS: Using the National Health Insurance Research database in Taiwan, we identified 2588 patients with T2DM who had developed severe hypoglycemia from 2001 to 2009. A comparison cohort who had never developed severe hypoglycemia was frequency matched at a ratio of approximately 1:2. Multivariate Cox proportional hazard regression analysis was used to evaluate the risk of hip fracture. RESULTS: During a median follow-up period of 3.9 years, there were 219 hip fracture events in 5173 comparison cohorts and 148 hip fracture events in 2588 hypoglycemia cohorts. The incidence of hip fracture was higher in patients with severe hypoglycemia than without severe hypoglycemia (17.19 vs. 8.83 per 1000 person-years; adjusted HR 1.71, 95% CI = 1.35-2.16). Approximately half of the individuals developed hip fracture within 2 years from the first occurrence of severe hypoglycemia. There was a significant associated trend towards increased hip fracture risk with increasing average visit of severe hypoglycemia per year (p for trend <0.001). Medication analysis showed that patients taking sulfonylurea alone, insulin alone, and insulin secretagogues combined with insulin had a higher associated risk to develop hip fracture. CONCLUSIONS: Severe hypoglycemia was associated with a higher risk to develop hip fracture. The more the visits of severe hypoglycemia per year indicated the higher associated risk in patients with T2DM. Fall is likely an important reason for severe hypoglycemia in relation to increased risk of hip fracture.

Fabrication of X-ray Microcalorimeter Focal Planes Composed of Two Distinct Pixel Types.

We are developing superconducting transition-edge sensor (TES) microcalorimeter focal planes for versatility in meeting specifications of X-ray imaging spectrometers including high count-rate, high energy resolution, and large field-of-view. In particular, a focal plane composed of two sub-arrays: one of fine-pitch, high count-rate devices and the other of slower, larger pixels with similar energy resolution, offers promise for the next generation of astrophysics instruments, such as the X-ray Integral Field Unit (X-IFU) instrument on the European Space Agency's Athena mission. We have based the sub-arrays of our current design on successful pixel designs that have been demonstrated separately. Pixels with an all gold X-ray absorber on 50 and 75 micron scales where the Mo/Au TES sits atop a thick metal heatsinking layer have shown high resolution and can accommodate high count-rates. The demonstrated larger pixels use a silicon nitride membrane for thermal isolation, thinner Au and an added bismuth layer in a 250 micron square absorber. To tune the parameters of each sub-array requires merging the fabrication processes of the two detector types. We present the fabrication process for dual production of different X-ray absorbers on the same substrate, thick Au on the small pixels and thinner Au with a Bi capping layer on the larger pixels to tune their heat capacities. The process requires multiple electroplating and etching steps, but the absorbers are defined in a single ion milling step. We demonstrate methods for integrating heatsinking of the two types of pixel into the same focal plane consistent with the requirements for each sub-array, including the limiting of thermal crosstalk. We also discuss fabrication process modifications for tuning the intrinsic transition temperature (Tc) of the bilayers for the different device types through variation of the bilayer thicknesses. The latest results on these "hybrid" arrays will be presented.

Second cytotoxic pathway of diphtheria toxin suggested by nuclease activity.

Diphtheria toxin (DTx) provokes extensive internucleosomal degradation of DNA before cell lysis. The possibility that DNA cleavage stems from direct chromosomal attack by intracellular toxin molecules was tested by in vitro assays for a DTx-associated nuclease activity. DTx incubated with DNA in solution or in a DNA-gel assay showed Ca2+- and Mg2+-stimulated nuclease activity. This activity proved susceptible to inhibition by specific antitoxin and migrated with fragment A of the toxin. Assays in which supercoiled double-stranded DNA was used revealed rapid endonucleolytic attack. Discovery of a DTx-associated nuclease activity lends support to the model that DTx-induced cell lysis is not a simple consequence of protein synthesis inhibition.

Fabrication of Flexible Superconducting Wiring with High Current-Carrying Capacity Indium Interconnects.

The X-ray integral field unit (X-IFU) is a cryogenic spectrometer for the Advanced Telescope for High ENergy Astrophysics (ATHENA). ATHENA is a planned next-generation space-based X-ray observatory with capabilities that surpass the spectral resolution of prior missions. Proposed device designs contain up to 3840 transition edge sensors, each acting as an individual pixel on the detector, presenting a unique challenge for wiring superconducting leads in the focal plane assembly. In prototypes that require direct wiring, the edges of X-IFU focal plane have hosted aluminum wirebonding pads; however, indium (In) 'bumps' deposited on an interface layer such as molybdenum nitride (MoN) can instead be used as an array of superconducting interconnects. We investigated bumped MoN:In structures with different process cleans and layer thicknesses. Measurements of the resistive transitions showed variation of transition temperature T c as a function of bias and generally differed from the expected bulk T c of In (3.41 K). Observed resistance of the In bump structures at temperatures below the MoN transition (at 8.0 K) also depended on the varied parameters. For our proposed X-IFU geometry (10 µm of In mated to a 1-µm In bump), we measured a minimum T c of 3.14 K at a bias current of 3 mA and a normal resistance of 0.59 mΩ per interconnect. We also investigated the design and fabrication of superconducting niobium (Nb) microstrip atop flexible polyimide. We present a process for integrating In bumps with the flexible Nb leads to enable high-density wiring for the ATHENA X-IFU focal plane.

Immunotoxicity of alcohol in young and old mice. II. Impaired T cell proliferation and T cell-dependent antibody responses of young and old mice fed ethanol-containing liquid diet.

The effect of extended ethanol consumption of young and old BALB/c mice on the proliferative response to Concanavalin A (Con A) and T cell-dependent antibody response of their spleen cells to sheep red blood cell (RBC) stimulation was determined. Splenic cells of young (3 months) and old (25 months) BALB/c mice, fed with one of three different diets (ethanol, maltose-substitute and standard mouse chow), were first cultured with Con A to assess T cell proliferation and production of interleukin 2 (IL2). Then, Con A-activated T blast cells from young and old mice were assessed for their proliferative responding capacity to exogenous human recombinant IL2 and crude rat IL2 supernatant. Finally, splenic cells of young and old mice were assessed for their ability to generate plaque-forming cells in response to sheep RBC. The results revealed that both T cell mitogenesis and IL2-dependent proliferation of T blast cells from young and old ethanol diet-fed mice were remarkably diminished as compared to that of young and old maltose-substituted diet (isocaloric control) fed mice, respectively. The ability of T cells from both young and old ethanol diet-fed mice to produce IL2, however, was not affected. Finally, the ability of young and old ethanol diet-fed mice to mount a primary antibody response to SRBC was also significantly reduced. These results taken together demonstrate for the first time that both T cell proliferative activity and T cell-dependent antibody response of young and old ethanol diet-fed mice are impaired; however, with respect to age, a differential effect of immunosuppression of ethanol was not noted.

Loss of hybridizable ribosomal DNA from human post-mitotic tissues during aging: II. Age-dependent loss in human cerebral cortex--hippocampal and somatosensory cortex comparison.

DNA was isolated from the hippocampal and from the somatosensory cortex of 13 humans (at autopsy). In both the cortex and hippocampus, the loss of ribosomal DNA (rDNA), as measured through hybridization in the liquid phase, approximates about 0.9% per year. The r value for somatosensory cortex was about -0.7 and that for the hippocampus was about -0.91. The correlation coefficient between the sets of two samples derived from the same individual (two different areas) in +0.945. These results are consistent with those reported concurrently for human myocardium and with earlier studies conducted with beagle dogs, in which only post-mitotic tissues (brain, heart and skeletal muscle) showed measurable decrements in these key genes. To the degree that the synthesis of new proteins is essential for sustained mental activity, these results are consistent with the observations that Nissl substance is more slowly replenished, following exhaustive work by motor cortical cells, and the fact that many older persons experience mental fatigue during continuous mental work at earlier times than do younger persons. The mechanism of loss is not certain, but may well be related to inadequacies in DNA repair systems, thereby allowing deletion of tandemly duplicated genes through cross-over "episome" formation, followed by degradation of the excised DNA segments. The ratio of loss of rDNA hybridizability in human and dogs in about 1 to 7, which approximates the relative ratios of their lifespans (reciprocals).

Requirement of 2-mercaptoethanol for in vitro growth factor production by T cells and vulnerability of the response to age.

The effects of 2-mercaptoethanol and age on spleen cells derived from young (3-4 months) and old (24-months) C57BL/6 mice were measured with respect to T cell growth factor or Interleukin 2 production. It was shown that: (A) 2-mercaptoethanol (or some homologue) is absolutely required for T cell growth factor production in vitro by murine cells (the optimum concentration is 5 X 10(-5) M for both young and old cells); (B) old cells are less responsive to suboptimum concentrations than young cells but their responses are not reduced to the same degree as young cells by supraoptimum toxic doses of 2-mercaptoethanol; and (C) at optimum 2-mercaptoethanol concentrations young and old cells have similar kinetic responses for T cell growth factor production and the accumulation of the T cell growth factor reaches a maximum between 18 and 24 hours. Considerations are presented of 2-mercaptoethanol (or some homologue) in its role as an important reactant in the production of T cell growth factor and in its susceptibility to aging.

Aging decreases beta-endorphin enhancement of T-cell mitogenesis in mice.

Numerous studies have demonstrated the role of the central nervous system in immunomodulation. beta-Endorphin, a neuropeptide that is released along with adrenocorticotropin by the pituitary in response to stress, has been shown to have various effects on immune function, although these effects are dependent on dose, animal model, and immune cell tested. Since the increased risk for infection and tumor that is observed in the elderly is thought to be in part secondary to waning cell-mediated immunity, we investigated the effect of age on beta-endorphin immunomodulation of T-cell proliferation in a murine model. Spleen cells obtained from young and old BALB/c mice were cultured in vitro with various mitogens with and without beta-endorphin. beta-Endorphin at 10(-8) M on day 3 of culture significantly enhanced concanavalin A (2.0 micrograms/10(6) cells per ml) mitogenesis but not phytohemagglutinin or lipopolysaccharide mitogenesis. Moreover, this enhancement was shown only in spleen cells from young mice and was not blocked by the opiate receptor antagonist naloxone, which suggests that enhancement of mitogenesis by beta-endorphin was mediated by a non-opiate receptor. Finally, our results support an altered response to neuroimmunomodulation with age.

Loss of hybridizable ribosomal DNA from human post-mitotic tissues during aging: I. Age-dependent loss in human myocardium.

DNA obtained from 29 male humans at autopsy was hybridized in liquid phase with tritium-labelled 18 and 28S ribosomal RNA (rRNA) in order to determine whether a change in the dosage in rDNA, which codes for rRNA, occurs during human aging. The individuals ranged in age from 3 months to 76 years. It was found that the amount of rDNA hybridizable per 260 nm absorption unit by DNA decreases by about 0.5% per year with a regression coefficient of about -0.83. These findings confirm earlier ones from this laboratory and indicate either a loss of these key genes during aging of humans or decreased hybridizability due to some other factor or factors. In any event, this degree of loss or inactivation of genes involving an all protein synthesis would seem to impair function of post-mitotic cells in response to maximal stress to about the same degree that function is lost in various human organ systems during aging, as defined by Shock and others.

Selective isolation of transiently transfected cells from a mammalian cell population with vectors expressing a membrane anchored single-chain antibody.

We present here a novel technology for the rapid selection of transiently transfected cells from total populations in culture. This system utilizes recombinant antibody technology to produce a 'molecular hook' by displaying a hapten-binding single-chain antibody (sFv) on the surface of transfected cells. Mammalian cell lines from several origins were transiently transfected with a plasmid (pHook-1) that encodes an sFv fused with a transmembrane anchor and found to express and display the functional hapten-binding sFv on their membranes. Transfected cells were selected from total populations in culture by virtue of their ability to bind to hapten-coated magnetic beads. Some cell lines were able to display sFv sufficient for selection as early as 2 h post-transfection. SK-BR-3 human breast carcinoma cells were co-transfected with pHook-1 and pCR31acZ (expresses beta-galactosidase), selected, and assayed for beta-galactosidase activity. The positive correlation between sFv and beta-galactosidase expression in these cells (95% of selected cells also expressed beta-galactosidase activity) suggests that pHook-1 will be useful in isolating cells co-expressing an exogenous gene of interest. Another vector was constructed in which a gene of interest may be expressed from the same plasmid as the sFv 'hook'. This construct (pHook-2) allows the selection of a homogenous population of cells expressing exogenous genes without co-transfection or the generation of stable transfectants. In experiments where the lacZ gene was co-expressed with the sFv 'hook' from this single plasmid, 100% of 293 human kidney cells and 100% of SK-BR-3 cells selected with antigen-coated magnetic beads stained positively for beta-galactosidase activity. We propose that this system will be a valuable tool for studying the acute and chronic effects of the expression of a variety of wild type and mutant proteins.

Recruitment of host CD8+ T cells by tumor-infiltrating lymphocytes and recombinant interleukin-2 during adoptive immunotherapy of cancer.

BACKGROUND: Previously we demonstrated that optimal doses of tumor-infiltrating lymphocytes (TIL) concomitant with recombinant interleukin-2 (rIL-2) effectively mediated complete tumor regression of murine 3-day pulmonary metastases. METHODS: In the present study we have investigated the contribution of the host immune response to the effectiveness of adoptive immunotherapy with TIL in combination with low-dose rIL-2. All experiments were performed in a murine pulmonary metastases model induced by intravenous injection of methylcholanthrene-induced sarcoma (MCA-105) cells into C57BL/6 mice. As a novel approach we used monoclonal antibody specific for CD4+ or CD8+ T cells to deplete the host of its T-cell subpopulations. RESULTS: Depletion of host CD8+ T cells 24 hours after tumor injection and 48 hours before TIL+rIL-2 treatment abrogated all antitumor activity of this type of immunotherapy and resulted in significant metastatic pulmonary disease (p < 0.001). In contrast, depletion of host CD4+ T cells did not alter the efficacy of TIL+rIL-2 treatment in tumor eradication. The loss of tumoricidal activity of TIL+rIL-2 treatment in a CD8+ T cell-depleted host could be overcome by adding back normal uneducated splenocytes 2 hours after TIL therapy (p < 0.001). In contrast, adding back CD8- CD4+ splenocytes to a CD8+ T cell-depleted host 2 hours after TIL+rIL-2 treatment resulted in significant pulmonary disease comparable to untreated animals. CONCLUSIONS: We conclude that the recruitment of host CD8+ T cells by adoptively transferred TIL+rIL-2 appears to be important for effective tumor eradication in this type of immunotherapy.

Reactivation of murine tumour-infiltrating lymphocytes with solid-phase anti-CD3 antibody: in vitro cytokine production is associated with in vivo efficacy.

Previously we described the use of solid-phase anti-CD3 monoclonal antibody (mAb) to stimulate murine tumour-infiltrating lymphocytes (TIL) and their subsequent expansion in recombinant interleukin 2 (rIL-2). In a pulmonary metastases model using the methylcholanthrene-induced sarcoma MCA-105 anti-CD3 activated TIL were capable of eradicating disease similar to TIL cultured in rIL-2 only. Here we extend these observations by characterizing the biological effects of sequential solid-phase anti-CD3 activation. TIL from MCA-105 tumour activated with solid-phase anti-CD3 on day 1 were reactivated on day 14, or day 26, or both and compared to TIL grown in rIL-2 only or TIL activated with anti-CD3 once on day 1. Reactivation enhanced in vitro proliferation 1.8- to 4-fold compared to TIL activated once with anti-CD3 (P < 0.05). In addition, the total lytic capacity of the cultures was enhanced after reactivation without changing the phenotype of the TIL cultures. Reactivation resulted in a greater in vivo efficacy when the TIL were administered within 72 h of reactivation. In contrast, TIL activated with anti-CD3 on day 1 and day 14 were least effective of all TIL cultures (P < 0.05). This correlated with in vitro cytokine production. The most effective TIL cultures in vivo produced 4- to 100-fold higher amounts of cytokines, especially interferon gamma (IFN gamma) and granulocyte macrophage colony stimulating factor (GM-CSF), than the other cultures. On the other hand, the least effective in vivo TIL culture, TIL activated with anti-CD3 on day 1 and 14, produced little or no cytokines. These data suggest that in vitro production of cytokines is indicative of in vivo efficacy of anti-CD3 activated TIL.

Generational differences in selenium status of women.

In this cross-sectional study of three generations of women, daughters (19-26 yr), mothers (40-58 yr) and maternal grandmothers (67-84 yr) from the same 10 families in central Ohio were studied to determine the effect of life-cycle differences, including estrogen status, on selenium status. Plasma and red blood cell (RBC) selenium and glutathione peroxidase (GPx) activities were determined and typical dietary selenium intakes were calculated from food-frequency questionnaires. Selenium status was lowest in the oldest generation. Plasma selenium of daughters and grandmothers were significantly lower than those of mothers, and plasma GPx and RBC selenium of grandmothers were also lower than those of the mothers. A positive correlation (r = 0.42, p < 0.04) was found between plasma estrogen and plasma selenium concentrations. Selenium intakes of all groups were adequate and no differences in selenium intakes were found among groups. The results of this study indicate that selenium status fluctuates during the female life cycle and is related to estrogen status.

Evaluation of perception of insulin therapy among Chinese patients with type 2 diabetes mellitus.

AIMS: To evaluate whether perception of insulin therapy differs between patients with type 2 diabetes treated with insulin and those treated with oral hypoglycaemic agents (OHAs), and to examine whether gender, education level, injection duration and mode of injection were associated with the patients' perception of insulin therapy. METHODS: The validated Chinese version of the Insulin Treatment Appraisal Scale (ITAS) was used to evaluate the perception of insulin therapy among 100 insulin-treated patients and 100 OHA-treated patients. The higher the total score, the more negative is the appraisal. RESULTS: The OHA-treated group had a higher mean total score (20 items), a higher mean total score for 16 negative items and a lower mean total score for four positive items than the insulin-treated group. The proportion of participants who rated the negative items as "agree" or "strongly agree" was significantly higher in the OHA-treated group than in the insulin-treated group. In addition, the proportion of participants who rated the four positive items as "agree" or "strongly agree" was lower in the OHA-treated group than in the insulin-treated group. Gender, education level, duration of insulin injection and mode of injection did not have a significant impact on perception of insulin therapy. CONCLUSION: Chinese type 2 diabetic patients taking OHAs had more negative beliefs and attitudes towards insulin therapy than patients being treated with insulin. This difference was not associated with either gender or education level. Furthermore, neither injection duration nor type of device was related to perception of insulin therapy.

Renal toxicity in patients with multiple myeloma receiving zoledronic acid vs. ibandronate: a retrospective medical records review.

AIMS: This retrospective study investigated the rates of renal impairment in patients with multiple myeloma treated with zoledronic acid and ibandronate. MATERIALS AND METHODS: We retrospectively reviewed medical records in a German oncology clinic, from May 2001 to December 2005. Creatinine measurements were analyzed from baseline (before zoledronic acid or ibandronate treatment) to last evaluation for each patient. A total of 84 patients were included. RESULTS: Zoledronic acid increased the risk of renal impairment by approximately 3-fold compared with ibandronate (renal impairment rates: zoledronic acid 37.7% vs. ibandronate 10.5%, relative risk [RR]=3.6, P=0.0029 serum creatinine [SCr]; 62.3% vs. 23.7%, RR=2.6, P=0.0001 glomerular filtration rate [GFR]). Ibandronate-treated patients switched from zoledronic acid had a significantly higher risk of renal impairment than patients receiving ibandronate monotherapy (zoledronic acid over ibandronate 39.1% vs. ibandronate monotherapy 6.7%, RR= 5.9, P=0.028 [SCr]; 65.2% vs 26.7%, RR=2.4, P=0.022 [GFR]). Multivariate analysis found significantly higher hazard ratios for zoledronic acid over ibandronate (SCr: Cox = 4.38, P=0.01; Andersen-Gill=8.22, P < 0.01; GFR: Cox = 4.31, P < 0.01; Andersen-Gill = 3.71, P < 0.01). CONCLUSIONS: Overall, this retrospective study suggests that multiple myeloma patients are more likely to experience renal impairment with zoledronic acid than with ibandronate. The risk of renal impairment increased if patients had received prior therapy with zoledronic acid.

Iterative interlacing error diffusion for synthesis of computer-generated holograms.

The iterative interlacing error-diffusion technique is a combination of the error-diffusion and the modified iterative interlacing techniques for synthesizing computer-generated holograms. The iterative interlacing error-diffusion technique leads to a dramatic improvement in the quality of reconstructed images, provided that the two constant parameters involved in iterations are chosen properly.

Mechanism of ethanol-mediated immunosuppression in mice: ethanol suppresses T-cell proliferation without affecting IL2 production and IL2 receptor expression.

The effect of extended ethanol consumption in young C57BL/6 mice on T-cell proliferation was studied. Splenic cells of young mice (3-4 months old), fed with one of three different liquid diets (5% ethanol, maltose-substitute, or standard liquid diet) for 28-38 days were cultured with plant lectins to assess T-cell proliferation and IL2 production. Expression of T-cell subset markers (CD4+/CD8+) was also determined. Then, Con A-activated T blast cells were assessed for their ability to express IL2 receptor (IL2R) and to respond to IL2. Finally, the proliferative response of splenic cells to PMA/ionomycin was assessed. The results showed that both lectin- and PMA/ionomycin-induced mitogenesis and IL2-dependent proliferation of T-cells from ethanol diet-fed mice were diminished as compared with that of maltose-substitute diet or standard liquid diet. However, the ability of T-cells from ethanol diet-fed mice to produce IL2 and to express IL2 R or CD4+/CD8+ subset markers was not affected. Furthermore, the magnitude of ethanol-mediated suppression of T-cell proliferation induced by PMA/ionomycin was comparable with that induced by Con A. These results taken together indicate that ethanol suppresses T-cell proliferation by interfering with events following the IL2-IL2R interaction. Therefore, it is likely that ethanol inhibits murine T-cell proliferation by selectively affecting the progression (IL2R-mediated events) rather than the initiation (mitogenic receptor-mediated events) of the cell cycle.