Modification of the sensitivity of glucose sensor implanted into subcutaneous tissue

The mechanism of reducing the glucose sensitivity of sensors implanted into the subcutaneous tissue of the normal rat was evaluated (n = 10) by comparing sensitivities observed in vitro and in vivo. In vivo sensitivity was significantly lower than that observed in vitro before implantation (p < 0.005). Most interestingly, in vitro sensitivity immediately after explanation did not differ from that in vivo and increased progressively during rinsing (p < 0.02 after 30 min). These results demonstrate that the reduction of in vivo sensitivity was not due to a local factor or factors but to a reversible alteration of the glucose sensor characteristics induced in vivo by some local factor(s). This suggests that modifications of the outer sensor membrane, the nature of which remains to be determined, may prevent this effect and resolve the problem.     

Effect of duodenal glucose infusion on blood glucose concentration in endotoxin-treated calves

The effect of endotoxemia on the intestinal absorption of glucose was evaluated in nine experiments performed on seven 3- to 5-week-old calves fitted with a duodenal cannula. An intraduodenal glucose load trial (infusion of 2 g glucose/kg b.w. as a 10% aqueous solution through the cannula over 60 min) was conducted in a group of 5 calves three times during the 4-day period: 48 h before and at 2 and 24 h after i.v. injection of E. coli 0111:B4 endotoxin (LPS) at a dose of 0.1 microgram/kg b.w. Control calves were treated similarly but instead of glucose they were infused intraduodenally with deionised water at a volume of 20 ml/kg b.w. In trial with glucose load performed 48 h before LPS administration, blood glucose concentration increased during the absorptive phase from 4.32 +/- 0.32 mmol/l to 11.45 +/- 0.87 mmol/l at 60 min and then decreased to a minimum value of 3.16 +/- 0.51 mmol/l at 240 min. During the initial phase of endotoxemia, blood glucose concentration did not change from baseline values in both groups of calves. Glucose concentration in control calves started to decrease at 165 min reaching a minimum value of 1.39 +/- 0.17 mmol/l at 210 min and then increased to 2.44 +/- 0.11 mmol/l at 480 min after LPS administration. The intraduodenal infusion of glucose at 2 h after LPS administration resulted in an increase in blood glucose concentration during the absorptive phase only in one calf. Blood glucose concentration in this calf increased between 30 and 90 min reaching a maximum value of 7.19 mmol/l at 60 min, and then decreased to a minimal value of 0.94 mmol/l at 180 min after glucose load. In the remaining four calves in this group, blood glucose concentration ranged from 3.89 +/- 0.37 mmol/l to 4.48 +/- 0.45 mmol/l up to 120 min, and then steadily decreased to a minimal value of 2.41 +/- 0.41 mmol/l at 300 min. In trial with glucose load performed 24 h after LPS administration, the rate of entry of glucose into the circulation during the absorptive phase was similar to that observed in the trial performed 48 h before LPS administration. In conclusion, these results indicate that endotoxemia impairs the intestinal absorption of glucose in calves. The magnitude of the absorption disturbance may vary in individual calves, and the inhibitory effect of LPS on the intestinal glucose absorption lasts less than 24 h.     

Initial evaluation of a 290-microm diameter subcutaneous glucose sensor: glucose monitoring with a biocompatible, flexible-wire, enzyme-based amperometric microsensor in diabetic and nondiabetic humans

Results of the initial clinical evaluation in 20 human subjects of a subcutaneously implanted microsensor-based amperometrically glycemia-monitoring system, carried out between April 1994 and June 1995, are reported. The system was based on the electrical connection ("wiring") of the reaction centers of glucose oxidase to a gold electrode and on elimination of the chemicals that interfere with glucose monitoring through their horseradish peroxidase-catalyzed oxidation by internally generated hydrogen peroxide. The sensor was finer than a 29-gauge needle and had no leachable components. Because of its high selectivity for glucose, the sensor output was virtually nil at zero glucose level. This enables prompt "one-point" in vivo calibration of the sensor with a single blood glucose sample. Microsensors were subcutaneously implanted in ten nondiabetic and ten insulin-dependent diabetes mellitus (IDDM) volunteers. All subjects underwent standard meal tests and intravenous glucose-tolerance tests (IVGTT) in addition to hourly plasma glucose measurements. The sensor signals were continuously recorded, and the glucose concentration estimates were derived by calibrating the sensor using a single blood sample (one-point calibration). Regression analysis revealed that the sensor-estimated glucose concentrations were linearly related to the plasma glucose concentrations (r2 = 0.75) over a wide glucose concentration range (2-28 mmol/L) (sensor estimate = plasma 0.96 + 0.26 mmol/L). The difference between the estimated and actual glucose concentration was -0.13+/-0.23 mmol/L [mean +/-95% confidence interval (CI), n = 546], and 95% of the estimates fell in clinically acceptable zones of the Clarke error grid. The sensing delay time was 10.4+/-2.3 min as measured by the IVGTT. The subjects reported no discomfort associated with wearing the sensors.     

Use of electroencephalography to detect Alzheimer''s disease in Down''s syndrome

Asthma is one of the most common chronic illnesses in childhood. Increases in hospitalization rates have occurred in several countries. The cumulative risk of asthma requiring medical attention was 11.7% for males and 7.0% for females aged 0-4 in Manitoba, Canada, for the cohort of children born in 1984/1985. The cumulative risk of hospitalization for males was nearly twice that of females (2.1% vs. 1.1%). Disease onset was most likely at age 1 year. The risk of rehospitalization or return physician visit for asthma increased significantly with the number of prior hospitalizations and physician visits, respectively, which may reflect both the persistence of asthma and the difficulty of developing an effective disease management strategy.     

Use of contrast-enhanced MR imaging to detect sacroiliitis in children

PURPOSE: To verify the diagnostic value of contrast-enhanced MR imaging compared with conventional radiography in the diagnosis of sacroiliitis in children. DESIGN AND PATIENTS: Radiography and MR imaging of the sacroiliac joints were performed in 185 children subdivided into the following groups according to the modified European Spondyloarthropathy (SpA) Study Group (ESSG) criteria: group 1, undifferentiated spondyloarthropathy (uSpA) (n=53, 94.5% HLA-B27+); group 2, differentiated SpA (n=45, 93.3% HLA-B27+); group 3, patients with no signs of SpA other than oligoarthritis (n=39, 92.3% HLA-B27+); group 4, HLA-B27+ controls with various other non-SpA diagnoses (n=22); and group 5, HLA-B27-controls with various other non-SpA diagnoses (n=26). Radiographs were evaluated on the basis of the modified New York criteria independently by three experienced radiologists masked to the clinical data. In a second step, the same radiologists independently evaluated the MR images without knowledge of the clinical data and radiographic findings using the recently published criteria developed by our group. These criteria allow differentiation of acute and chronic inflammatory changes. RESULTS: Radiography demonstrated sacroiliitis in 18 patients: 4 of 53 in group 1 (7.5%), 14 of 45 in group 2 (31%), but none in groups 3, 4 and 5. In contrast, MR imaging demonstrated acute and/or chronic sacroiliitis in 44 patients: 18 of 53 in group 1 (34%), 21 of 45 in group 2 (46.7%) and 5 of 39 in group 3 (12.8%), but none in groups 4 and 5. The percentage of sacroiliitis detected by MR imaging was significantly higher than that detected by radiography (P<0.001). CONCLUSION: Contrast-enhanced MR imaging is a useful method for detecting sacroiliitis in children. Advantages of contrast-enhanced MR imaging compared with conventional radiography are a higher sensitivity due to the ability to document early and acute changes and the absence of radiation exposure.     

Use of a gastric juice-based PCR assay to detect Helicobacter pylori infection in culture-negative patients

A gastric juice-based PCR assay was compared with culture, microscopy, and a rapid urease test with specimens from 114 subjects. The PCR and conventional tests were positive for 76 and 62% of the subjects, respectively. The prevalence of gastroduodenal disease and seropositivity for anti-Helicobacter pylori immunoglobulin G were similarly high among conventional-test-positive and PCR-only-positive subjects compared to all-negative ones. The PCR assay is recommended to confirm the H. pylori status of culture-negative peptic-ulcer patients.     

Volume of blood required to detect common neonatal pathogens

OBJECTIVE: To determine the minimum volume of blood and the absolute number of organisms required for detection of bacteremia and fungemia by an automated colorimetric blood culture system (BacT/Alert, Organon Teknika). DESIGN: Common neonatal pathogens, Escherichia coli, Streptococcus agalactiae (group B streptococcus (GBS): one American Type Culture Collection (ATCC) strain and one clinical isolate), Staphylococcus epidermidis, and Candida albicans, were seeded into blood to produce bacteremia or fungemia with low colony counts (1 to 3 colony-forming units (CFU) per milliliter) and ultra-low colony counts (<1 CFU/ml). For each organism, 96 culture bottles were inoculated with either 0.25, 0.5, 1.0, or 4.0 ml of the two seeded blood concentrations. Blood culture bottles were incubated in the BacT/Alert device for 5 days, and time to positivity was noted when applicable. All bottles were subcultured on plated media. DATA ANALYSIS: The Poisson statistic was used to calculate the probability of finding at least one viable CFU per inoculated culture bottle. The fraction of culture bottles with positive findings per group was divided by the probability of one or more organisms present to give the positivity index. RESULTS: Plated subculture identified no growth of organisms not detected by the colorimetric detection system. The false-positive rate for the automated device was less than 1%. The positivity index for the GBS clinical isolate was 1.13, for the GBS ATCC isolate 0.96, for S. epidermidis 0.94, for C. albicans 0.97, and for E. coli 0.95. There was a statistically significant difference with time to positivity and inocula volume (p <0.01), but the difference was not clinically important. CONCLUSIONS: If one or two viable colony-forming units are in the blood inoculated into culture media, the BacT/Alert system will detect growth rapidly. Because there appears to be a sizable subset of neonates who are at risk of sepsis with a colony count less than 4 CFU/ml, then a 0.5 ml inoculum of blood into the culture media is inadequate for sensitive and timely detection of bacteremia. One to two milliliters of blood should increase microorganism recovery in the face of low-colony-count sepsis.     

Use of fluorescent in situ hybridization to detect aneuploidy in cervical dysplasia

An 8 year old girl with acute disseminated encephalomyelitis (ADEM) is described. Elevated serum antibody titers suggested recent Mycoplasma pneumoniae infection. T2-weighted image of magnetic resonance imaging (MRI) disclosed multiple lesions of high signal intensity in bilateral basal ganglia and thalami as well as in the white matter. Postcontrast T1-weighted image revealed an enhanced lesion in the deep white matter. She showed rapid clinical improvement in response to corticosteroid therapy. The lesions had disappeared completely on MRI performed 10 weeks after the onset. ADEM is believed to be a demyelinating disorder of probable autoimmune etiology. MRI findings in this case may support the hypothesis that the primary pathological event is vascular injury and demyelination occurs only as a secondary phenomenon.     

Use of a subcutaneous pedicle ulnar flap to cover skin defects around the wrist

6-thioguanine (6-TGN) and methyl 6-mercaptopurine nucleotides (Me6-MPNs) are the two major metabolites found in erythrocytes after administration of azathioprine. In an attempt to understand the role of these metabolites in the pharmacologic and toxic activity of thiopurines, we have developed a HPLC method for the simultaneous determination of 6-TGNs and Me6-MPNs in erythrocytes. A simple and rapid treatment procedure based on deproteinization by perchloric acid with dithiothreitol is described. The nucleotides were hydrolyzed to their own bases by heating the sample for 45 min at 100 degrees C. During acid hydrolysis Me6-MP was converted into a compound analyzed on a Purospher RP18-e column with 0.02 mol/L dihydrogenophosphate buffer-methanol as eluents. With this procedure, mean recoveries of 73.1% and 84.0% for 6-TGN and Me6-MPN derivatives, respectively, were found.     

Use of lambda phage DNA as a hybrid internal control in a PCR-enzyme immunoassay to detect Chlamydia pneumoniae

An inherent problem in the diagnostic PCR assay is the presence of ill-defined inhibitors of amplification which may cause false-negative results. Addition of an amplifiable fragment of foreign DNA in the PCR to serve as a hybrid internal control (HIC) would allow for a simple way to identify specimens containing inhibitors. Two oligonucleotide hybrid primers were synthesized to contain nucleic acid sequences of the Chlamydia pneumoniae 16S rRNA primers in a position flanking two primers that target the sequences of a 650-bp lambda phage DNA segment. By using the hybrid primers, hybrid DNA comprising a large sequence of lambda phage DNA flanked by short pieces of chlamydia DNA was subsequently generated by PCR, cloned into a plasmid vector, and purified. Plasmids containing the hybrid DNA were diluted and used as a HIC by adding them to each C. pneumoniae PCR test. Consequently, C. pneumoniae primers were able to amplify both chlamydia DNA and the HIC DNA. The production of a 689-bp HIC DNA band on an acrylamide gel indicated that the specimen contained no inhibitors and that internal conditions were compatible with PCR. Subsequently, a biotinylated RNA probe for the HIC was transcribed from a nested sequence of the HIC and was used for its hybridization. Detection of the HIC DNA-RNA hybrid was achieved by enzyme immunoassay (EIA). This PCR-EIA system with a HIC was initially tested with 12 previously PCR-positive and 14 previously PCR-negative specimens. Of the 12 PCR-positive specimens, 11 were reconfirmed as positive; 1 had a negative HIC value, indicating inhibition. Of the 14 previously PCR-negative specimens, 13 were confirmed as true negative; 1 had a negative HIC value, indicating inhibition. The assay was then used with 237 nasopharyngeal specimens from patients with pneumonia. Twenty-one of 237 (8.9%) were positive for C. pneumoniae, and 42 (17.7%) were found to inhibit the PCR. Specimens showing inhibitory activity were diluted 1:10 and were retested. Ten specimens were still inhibitory to the PCR and required further DNA purification. No additional positive samples were detected and 3 nasopharyngeal specimens remained inhibitory to PCR. Coamplification of a HIC DNA can help confirm true-negative PCR results by ruling out the presence of inhibitors of DNA amplification.     

Long-term observation of subcutaneous tissue reaction to synthetic auditory ossicle (Apaceram) in rats

The present study evaluates histological characteristics of the soft tissue response to long-term implantation of Apaceram discs composed of dense hydroxyapatite in rats. Discs were implanted into the subcutaneous tissue of 76 rats for six to 20 months. Decalcified histological sections stained with haematoxylin and eosin (H & E) and Mallory's azan were examined. Different cell types surrounding implants were counted. The greatest proportion of macrophages was found at six months (13.5 per cent). This proportion gradually decreased to four per cent at 20 months. Small numbers of lymphocytes and foreign body giant cells were observed in every group, but neither neutrophils nor osteogenesis were observed in any specimens. Results of the present study and previous related studies indicate that despite reappearance of a small number of macrophages six months after implantation, Apaceram is useful for reconstructive surgery.     

A new enzyme immunoassay to detect antibodies to arboviruses in the blood of wild birds

A selective recruitment of eosinophils to sites of inflammation is claimed to be controlled by regulation of cytokines, chemokines and adhesion molecules. In animal models, eotaxin has been suggested to be a potent chemokine since it in cooperation with interleukin-5 induce selective chemotaxis and infiltration of eosinophils to lung tissue after an allergen provocation. We have investigated the in vitro effect of eotaxin on human peripheral blood eosinophils with respect to CD11b/CD18 expression and adhesion properties to the matrix protein fibronectin. We did not find any effect of eotaxin per se on CD11b/CD18 expression, neither on eosinophils from healthy subjects nor from patients with asymptomatic pollen related asthma. However, eotaxin significantly upregulated the quantitative level of CD11b/CD18 and increased the adhesion to fibronectin in eosinophils from healthy subjects preincubated in vitro with interleukin-5, but not in eosinophils preincubated with fMLP. Moreover, eosinophils harvested 24 hours after an in vivo allergen inhalation provocation in asthmatics, upregulated CD11b/CD18 after in vitro incubation with eotaxin alone.     

Removal of adenosine decreases the responsiveness of muscle glucose transport to insulin and contractions

Adenosine in the extracellular space modulates stimulated glucose transport in striated muscle. In the heart and in adipocytes, adenosine potentiates insulin-stimulated glucose transport. There is controversy regarding the effect of adenosine in skeletal muscle, with reports of both an inhibitory effect and no effect, on insulin-stimulated glucose transport. We found that, in rat epitrochlearis and soleus muscles, removing adenosine with adenosine deaminase or blocking its action with the adenosine receptor blocker CPDPX markedly reduces the responsiveness of glucose transport to stimulation by 1) insulin alone, 2) contractions alone, and 3) insulin and contractions in combination. Measurement of the increase in GLUT4 at the cell surface in response to a maximally effective insulin stimulus in the epitrochlearis muscle, using the exofacial label ATB-[3H]BMPA, showed that adenosine deaminase treatment markedly reduces cell-surface GLUT4 labeling. The reduction in cell-surface GLUT4 labeling was similar in magnitude to the decrease in maximally insulin-stimulated glucose transport activity in adenosine deaminase-treated muscles. These results show that adenosine potentiates insulin- and contraction-stimulated glucose transport in skeletal muscle by enhancing the increase in GLUT4 at the cell surface and raise the possibility that decreased adenosine production or action could play a causative role in insulin resistance.     

Intracellular glucose concentration in derepressed yeast cells consuming glucose is high enough to reduce the glucose transport rate by 50%

1. This study describes the in vitro characterization of two potent and selective 5-HT6 receptor antagonists at the rat and human recombinant 5-HT6 receptor. 2. In binding assays with [3H]-LSD, 4-amino-N-(2,6 bis-methylamino-pyrimidin-4-yl)-benzene sulphonamide (Ro 04-6790) and 4-amino-N-(2,6 bis-methylamino-pyridin-4-yl)-benzene sulphonamide (Ro 63-0563) had mean pKi values +/-s.e.mean at the rat 5-HT6 receptor of 7.35+/-0.04 and 7.83+/-0.01, respectively and pKi values at the human 5-HT6 receptor of 7.26+/-0.06 and 7.91+/-0.02, respectively. 3 .Both compounds were found to be over 100 fold selective for the 5-HT6 receptor compared to 23 (Ro 04-6790) and 69 (Ro 63-0563) other receptor binding sites. 4. In functional studies, neither compound had any significant effect on basal levels of cyclicAMP accumulation in Hela cells stably expressing the human 5-HT6 receptor, suggesting that the compounds are neither agonists nor inverse agonists at the 5-HT6 receptor. However, both Ro 04-6790 and Ro 63-0563 behaved as competitive antagonists with mean +/-s.e.mean pA2 values of 6.75+/-0.07 and 7.10+/-0.09, respectively. 5. In rats habituated to observation cages, Ro 04-6790 produced a behavioural syndrome similar to that seen following treatment with antisense oligonucleotides designed to reduce the expression of 5-HT6 receptors. This behavioural syndrome consisted of stretching, yawning and chewing. 6. Ro 04-6790 and Ro 63-0563 represent valuable pharmacological tools for the identification of 5-HT6 receptors in natural tissues and the study of their physiological function.     

Acrophase amplitude of ambulatory blood pressure decreases in migraineurs

Ambulatory blood pressure was recorded and analyzed in 13 migraineurs during headache-free periods and in 11 healthy subjects. Systolic blood pressure, mean blood pressure, diastolic blood pressure, pulse pressure, and pulse rate were recorded for 48 hours (three times every hour from 6 AM to mid-night, once an hour at night). Circadian variation of blood pressure was analyzed using single cosinor analysis and group mean cosinor analysis methods. Single cosinor analysis identified significant circadian rhythm of systolic blood pressure in 10 of 11 control subjects (90.9%) and 5 of 13 migraineurs (38.5%, P < 0.02 versus controls, Fisher's exact test). Incidences of significant circadian rhythm of mean blood pressure and diastolic blood pressure were 100% and 100% in controls, 46.2% and 53.8% in the migraineurs (P < 0.01, P < 0.02 versus controls). Incidences of significant rhythm of pulse pressure were 36.3% in controls and 38.5% in the migraine group (difference was not significant). Group mean cosinor analysis identified significant circadian rhythm in both the migraine group and the controls. The MESOR (midline estimating statistic of rhythm) values of systolic, mean, and diastolic blood pressures showed no significant differences between the migraine group and the controls. Acrophase amplitudes of systolic, mean, and diastolic blood pressure were 4.2, 5.2, and 5.9 mm Hg in the migraine group, respectively; and 7.2, 7.3, and 7.5 mm Hg in the controls, respectively. These amplitudes of systolic, mean, and diastolic blood pressures in the migraine group were significantly smaller than those in the controls. These data suggest that some migraineurs lose or alter their circadian blood pressure rhythm. Evaluating migraineurs as a group, significant circadian rhythm of blood pressure can be identified and oscillation amplitudes of blood pressures are decreased. The present results suggest that migraineurs may be subject to dysfunction of the circadian rhythm generator and the autonomic nervous system. Possible involvement of serotonergic projections from the raphe to the suprachiasmatic nuclei of the hypothalamus in migraine is discussed.     

Effects of diet enrichment with glucose and casein on blood cortisol concentration of calves in early postnatal period

The studies were carried out on 30 calves from the 4th to the 25th day of life, divided in three groups; control, glucose and casein. The glucose group was fed a diet containing 25% more glucose than the control. The increased dietary glucose supply decreased cortisol concentration in blood serum. The casein group was fed a diet containing 25% more protein-casein-than the control. The increased dietary casein supply increased the blood serum cortisol concentration. The changes discussed were observed not only during the period of diet application, but also a few days following the termination of the diet.