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PubMed1996

Insulating the scrotal neck affects semen quality and scrotal/testicular temperatures in the bull.

Nine Simmental X Angus bulls (2-yr of age) were used in 2 experiments. In Experiment 1, the scrotal neck was insulated (from Day 1 to Day 8) in 5 bulls, and semen was collected from all 9 bulls by electroejaculation approximately every 3 d until Day 35. Bulls with insulated scrotal necks had lower percentages of normal spermatozoa (P < 0.08) and higher percentages of spermatozoa with head defects (P < 0.06) or droplets (P < 0.08) than the untreated bulls. There was a time-by-treatment interaction (P < 0.04) for midpiece defects; the incidence was higher (P < 0.05) in the insulated than noninsulated bulls from Day 5 to Day 32. Spermatozoa within the epididymis or at the acrosome phase during insulation appeared to be the most affected. Compared with the noninsulated bulls, the insulated bulls had twice as many (P < 0.02) spermatozoa with midpiece defects and 4 times as many (not significant) with droplets on Day 5, fewer (P < 0.04) normal spermatozoa and 3 times as many with midpiece defects (P < 0.05) and with droplets (not significant) on Day 8, fewer (P < 0.02) normal spermatozoa on Days 15 and 18, and more sperm cells (P < 0.05) with head defects on Days 18 and 21. In Experiment 2, scrotal subcutaneous temperature (SQT; degrees C, mean +/- SE) prior to and after the scrotal neck had been insulated for 48 h in all 9 bulls was 30.4 +/- 0.7 and 32.4 +/- 0.6 (P < 0.01) at the top, 30.3 +/- 0.7 and 31.8 +/- 0.6 (P < 0.03) at the middle, and 30.2 +/- 0.8 and 30.7 +/- 0.6 (P < 0.05) at the bottom of the scrotum. Concurrently, there was an increase (0.9 degrees C) in intratesticular temperature (ITT) at the top (P < 0.07), middle (P < 0.04), and bottom (P < 0.04) of the testes. Scrotal surface temperature (SST) prior to and after the scrotal neck had been insulated for 24 h was 29.2 +/- 0.7 and 28.2 +/- 0.4 (P < 0.05) at the top of the scrotum and 24.7 +/- 0.6 and 25.3 +/- 0.7 (not significant) at the bottom, resulting in SST gradients of 4.6 +/- 0.6 and 2.9 +/- 0.5, respectively (P < 0.05). However, after the scrotal neck had been insulated for 48 h, none of the SST end points were significantly different from those prior to insulation. It appears that compensatory thermoregulatory mechanisms restored SST but were not able to restore SQT and ITT. Insulation of the scrotal neck affected SST, SQT, ITT and semen quality, emphasizing the importance of the scrotal neck in scrotal/testicular thermoregulation.

Journal Article↗

PubMed2004

Direct comparison of the insulating properties of two genetic elements in an adenoviral vector containing two different expression cassettes.

Targeted gene expression can be achieved through the use of cell-selective promoters. However, when the expression cassette is delivered by an adenovirus, "promoter interference," resulting in the loss of specificity, has been reported. To overcome this problem, insulator elements (the bovine growth hormone transcription stop signal or HS4 chromatin insulators of the chicken beta-globin locus) have been used. The present study examines the efficacy of these insulators elements, when two independent expression cassettes (one in which a strong, ubiquitous promoter drives the expression of the green fluorescent protein and another in which the "cancer-selective" ERBB2 promoter drives the expression of the herpes simplex virus thymidine kinase [HSVtk] gene) are incorporated into the same recombinant adenovirus. As expected, the presence of either insulator does not alter the expression of HSVtk in ERBB2-positive cells, measured through sensitization of the cells to ganciclovir. When ERBB2-negative cells were infected at a multiplicity of infection (MOI) of 10, the uninsulated virus sensitized ERBB2-negative cells to the same extent as it did for ERBB2-positive cells; partial sensitization was observed when transcriptional terminators were used, indicating a partial insulating effect; and complete insulation (no sensitization to ganciclovir) was observed when HS4 chromatin insulators were used. However, this complete insulation was lost when the MOI of virus was increased to 100. Our study demonstrates the possibility of insulating a conditionally expressed transgene in the vicinity of another expression cassette, but this insulating effect is lost when the multiplicity of infection is increased.

Adenoviridae↗

PubMed2000

Differences in insulator properties revealed by enhancer blocking assays on episomes.

Insulators are genomic elements that define domains of transcriptional autonomy. Although a large number of insulators have been isolated, it is unclear whether these elements function by shared molecular mechanisms. Novel applications of FLP recombinase technology were used to dissect and compare the function of the Drosophila: gypsy and scs insulators. Inter actions between FLP monomers bound to chromosomally integrated FRT sites were unimpeded by either insulator, demonstrating that these insulators do not establish a chromosomal environment capable of disrupting all types of protein-protein interactions. The gypsy insulator blocked enhancer-activated transcription on FLP-generated extra-chromosomal episomes, whereas the scs insulator displayed silencing effects. These data indicate that these insulators differ in the mechanisms used to prevent enhancer function. That the gypsy insulator blocked enhancer-promoter communication within small episomes suggests that these effects may be accomplished without a global reorganization of chromatin structure. Instead, the gypsy insulator may disrupt enhancer-activated transcription by direct interference with transmission of the enhancer signal to the promoter.

Animals↗

PubMed2001

The gypsy insulator of Drosophila affects chromatin structure in a directional manner.

Chromatin insulators are thought to regulate gene expression by establishing higher-order domains of chromatin organization, although the specific mechanisms by which these sequences affect enhancer-promoter interactions are not well understood. Here we show that the gypsy insulator of Drosophila can affect chromatin structure. The insulator itself contains several DNase I hypersensitive sites whose occurrence is dependent on the binding of the Suppressor of Hairy-wing [Su(Hw)] protein. The presence of the insulator in the 5' region of the yellow gene increases the accessibility of the DNA to nucleases in the promoter-proximal, but not the promoter-distal, region. This increase in accessibility is not due to alterations in the primary chromatin fiber, because the number and position of the nucleosomes appears to be the same in the presence or absence of the insulator. Binding of the Su(Hw) protein to insulator DNA is not sufficient to induce changes in chromatin accessibility, and two domains of this protein, presumed to be involved in interactions with other insulator components, are essential for this effect. The presence of Modifier of mdg4 [Mod(mdg4)] protein, a second component of the gypsy insulator, is required to induce these alterations in chromatin accessibility. The results suggest that the gypsy insulator affects chromatin structure and offer insights into the mechanisms by which insulators affect enhancer-promoter interactions.

Animals↗

PubMed1996

Sudden infant death syndrome: insulation from bedding and clothing and its effect modifiers. The National Cot Death Study Group.

BACKGROUND: Thermal stress related to excessive insulation from bedding and clothing has been postulated to be associated with sudden infant death syndrome (SIDS). METHODS: The parents of 393 (81%) of the infants who died of SIDS in the post neonatal period were interviewed at home. Interviews were also completed with the parents of 1592 (88.4% of total) controls, a representative sample of all hospital births. The study was conducted in regions of the North and South Island of New Zealand in which 78% of all New Zealand births occurred in 1987-1990. Temperatures for the infant's bedroom estimated from the outside temperature and a model were used to predict the appropriate insulation for lower critical temperature (temperature below which the metabolic rate is likely to increase). RESULTS: Sudden infant death syndrome was associated with extra thermal insulation of > 2 tog above the lower critical value, the odds ratio (OR) was 1.70 (95% confidence interval [Cl] : 1.3-2.20) after adjusting for season. After adjusting for a number of confounding factors the CR was reduced to 1.35 (95% Cl: 0.97-1.87). Also associated with SIDS was too little thermal insulation OR = 1.67(95% Cl: 1.13-2.48; and 2.63 (95% Cl: 1.61-4.30) when adjustments were made for the confounding factors. The interaction effect between infants sleeping prone and >2 tog extra thermal insulation was significant (OR = 6.07, 95% Cl: 3.83-9.60). Infants with too little thermal insulation were at increased risk if they were not tightly wrapped (OR = 3.81, 95% Cl: 2.04-7.09). There were also small additive interaction effects if the mother smoked and the infants had > 2 tog extra thermal insulation, or they were ill and had > 4 tog extra insulation. Interaction effects between thermal insulation and other factors were not significant. CONCLUSION: More thermal insulation than was necessary to maintain the lower critical temperature increased the risk of SIDS primarily among infants in the prone sleep position, and to a lesser degree in infants whose mothers smoked and in infants who were unwell.

Bedding and Linens↗

PubMed1999

Change of footwear insulation at various sweating rates.

Moisture inside the footwear can considerably affect the thermal insulation. In this study with a thermal foot model there was simulated three sweat rates (3, 5 and 10 g/h). Five types of footwear with various insulation levels (dry insulation from 0.19 to 0.50 m2. K/W) were tested. The footwear insulation reduction was calculated for 1.5 hour period. The reduction in insulation was related to sweating rate and initial insulation. The footwear with high insulation lost even in percentile more insulation than thin boots under the same conditions (9-19% at 3 g/h, 13-27% at 5 g/h and 19-36% at 10 g/h). A relationship between insulation decrease and sweating rate was established. An 8-hour sweating test (5 g/h) and a test for determining evaporative heat, losses were carried out in addition. The insulation reduction during the first 1.5 hours of the 8-hour test answered for more than half of the total reduction.

Body Temperature Regulation↗

PubMed1989

Recurrence of insulitis in the NOD mouse after early prolonged anti-CD4 monoclonal antibody treatment.

The treatment of young NOD mice with continuous anti-CD4 mAb administration has previously been shown to prevent insulitis from developing. It has also been shown that insulitis, once present, can be abolished by anti-CD4 mAb treatment, but that the insulitis recurs after treatment stops. We have studied the effect of treating young NOD/Wehi mice with a limited course of anti-CD4 mAb to determine if any long term prevention of insulitis ensued. Anti-CD4 mAb (H129.19) was given i.p. weekly to 10 female NOD/Wehi mice from 25-100 days of age. Control mice received no treatment. The pancreas of the mice were examined histologically at 100 and 180 days of age and severity of insulitis graded from 0-100%. The insulitis grade in treated mice at 100 days was 5 +/- 2% compared with 29 +/- 26% in age matched control mice (p less than 0.01). At 180 days of age the insulitis grade in treated mice was 29 +/- 23% compared with 47 +/- 22% in aged matched controls (NS). Thus insulitis was prevented from developing by early treatment with anti-CD4 mAb but developed later when treatment was stopped. Early anti-CD4 mAb treatment does not therefore appear to produce a tolerant state so averting insulitis development and, insulitis can still develop in older NOD mice.

Animals↗

PubMed1984

Decrease in body insulation with exercise in cool water.

Steady-state body insulation was measured in 7 healthy male subjects during rest and exercise for 3 h in water of 28 degrees C - 32 degrees C. At rest, maximal body insulation increased as a linear function of mean subcutaneous fat thickness by an amount approximately 4-fold what would be predicted from the physical insulation of fat alone. With arm plus leg exercise, body insulation declined as an exponential function of the exercise intensity, reaching approximately 25% of the resting value at work loads above VO2 = 1.2 liters.min-1. During exercise the relationship between overall body insulation and mean subcutaneous fat thickness was almost identical to that predicted from fat insulation and mean subcutaneous fat thickness was almost identical to that predicted from fat insulation alone. These results suggest that 75% of maximal body insulation in resting subjects is achieved by use of skeletal muscle as an insulative barrier and that the muscle component is increased with increasing fat thickness. This muscle insulation shell is lost during exercise. As a practical consequence, heat generated by muscular exercise in water colder than critical water temperature cannot offset cooling unless the exercise intensity is great.

Adipose Tissue↗

PubMed2001

Biodegradation of polyether polyurethane inner insulation in bipolar pacemaker leads.

Several bipolar coaxial pacemaker leads, composed of an outer silicone rubber insulation and an inner polyether polyurethane (PEU) insulation, which were explanted due to clinical evidence of electrical dysfunction, were analyzed in this study. Optical microscopy (OM) and scanning electron microscopy (SEM) were used to determine the cause of failure. Attenuated total reflectance-Fourier transform infrared microscopy (ATR-FTIR) was used to analyze the PEU insulation for chemical degradation. In all leads, the silicone rubber outer insulation showed no signs of physical damage. Physical damage to the inner PEU insulation was the source of electrical dysfunction. Cracks through the PEU compromised the insulation between the inner and outer conductor coils in the lead. It was observed with SEM that these cracks originated on the outer surface of the inner insulation and progressed inward. ATR-FTIR analysis showed that the PEU had chemically degraded via oxidation of the ether soft segment. Furthermore, it was revealed that chemical degradation was more advanced on the outer surface of the PEU. It was hypothesized that hydrogen peroxide permeated through the outer silicone insulation and decomposed into hydroxyl radicals that caused the chemical degradation of PEU. The metal in the outer conductor coil catalyzed the decomposition of the hydrogen peroxide. Chemical degradation of the PEU could also have been catalyzed by metal ions created from the corrosion of the metal in the outer conductor coil by hydrogen peroxide. Physical damage probably occurred in regions of the leads that were subjected to a higher hydrogen peroxide concentration from inflammatory cells and high degrees and rates of strain due to intercorporeal movement, including, but not limited to, cardiac movement. Chemical degradation and physical damage probably had a synergistic affect on failure of the insulation, in that as chemical degradation proceeded, the polymer surface became brittle and more susceptible to physical damage. As physical damage proceeded, cracks propagated into the unaffected bulk, exposing it to oxidants.

Biocompatible Materials↗

PubMed2006

3D quantification of microclimate volume in layered clothing for the prediction of clothing insulation.

Garment fit and resultant air volume is a crucial factor in thermal insulation, and yet, it has been difficult to quantify the air volume of clothing microclimate and relate it to the thermal insulation value just using the information on the size of clothing pattern without actual 3D volume measurement in wear condition. As earlier methods for the computation of air volume in clothing microclimate, vacuum over suit and circumference model have been used. However, these methods have inevitable disadvantages in terms of cost or accuracy due to the limitations of measurement equipment. In this paper, the phase-shifting moiré topography was introduced as one of the 3D scanning tools to measure the air volume of clothing microclimate quantitatively. The purpose of this research is to adopt a non-contact image scanning technology, phase-shifting moiré topography, to ascertain relationship between air volume and insulation value of layered clothing systems in wear situations where the 2D fabric creates new conditions in 3D spaces. The insulation of vests over shirts as a layered clothing system was measured with a thermal manikin in the environmental condition of 20 degrees C, 65% RH and air velocity of 0.79 m/s. As the pattern size increased, the insulation of the clothing system was increased. But beyond a certain limit, the insulation started to decrease due to convection and ventilation, which is more apparent when only the vest was worn over the torso of manikin. The relationship between clothing air volume and insulation was difficult to predict with a single vest due to the extreme openings which induced active ventilation. But when the vest was worn over the shirt, the effects of thickness of the fabrics on insulation were less pronounced compared with that of air volume. In conclusion, phase-shifting moiré topography was one of the efficient and accurate ways of quantifying air volume and its distribution across the clothing microclimate. It is also noted that air volume becomes more crucial factor in predicting thermal insulation when clothing is layered.

Body Temperature Regulation↗

PubMed2004

The centrosomal protein CP190 is a component of the gypsy chromatin insulator.

Chromatin insulators, or boundary elements, affect promoter-enhancer interactions and buffer transgenes from position effects. The gypsy insulator of Drosophila is bound by a protein complex with two characterized components, the zinc finger protein Suppressor of Hairy-wing [Su(Hw)] and Mod(mdg4)2.2, which is one of the multiple spliced variants encoded by the modifier of mdg4 [mod(mdg4)] gene. A genetic screen for dominant enhancers of the mod(mdg4) phenotype identified the Centrosomal Protein 190 (CP190) as an essential constituent of the gypsy insulator. The function of the centrosome is not affected in CP190 mutants whereas gypsy insulator activity is impaired. CP190 associates physically with both Su(Hw) and Mod(mdg4)2.2 and colocalizes with both proteins on polytene chromosomes. CP190 does not interact directly with insulator sequences present in the gypsy retrotransposon but binds to a previously characterized endogenous insulator, and it is necessary for the formation of insulator bodies. The results suggest that endogenous gypsy insulators contain binding sites for CP190, which is essential for insulator function, and may or may not contain binding sites for Su(Hw) and Mod(mdg4)2.2.

Alleles↗

PubMed1998

A nuclear matrix/scaffold attachment region co-localizes with the gypsy retrotransposon insulator sequence.

The 5'-untranslated region of the Drosophila gypsy retrotransposon contains an "insulator," which disrupts the interactions between enhancer and promoter elements located apart. The insulator effect is dependent on the suppressor of Hairy-wing (su(Hw)) protein, which binds to reiterated sites within the 350 base pairs of the gypsy insulator, whereby it additionally acts as a transcriptional activator of gypsy. Here, we show that the 350-base pair su(Hw) binding site-containing gypsy insulator behaves in addition as a matrix/scaffold attachment region (MAR/SAR), involved in interactions with the nuclear matrix. In vitro experiments using nuclear matrices from Drosophila, murine, and human cells demonstrate specific binding of the gypsy insulator, not observed with any other sequence within the retrotransposon. Moreover, we show that the gypsy insulator, like previously characterized MAR/SARs, specifically interacts with topoisomerase II and histone H1, i.e. with two essential components of the nuclear matrix. Finally, experiments within cells in culture demonstrate differential effects of the gypsy MAR sequence on reporter genes, namely no effect under conditions of transient transfection and a repressing effect in stable transformants, as expected for a sequence involved in chromatin structure and organization. A model for the gypsy insulator, which combines within a short "compacted" retroviral sequence three functional domains (insulator, enhancer, and the presently unraveled MAR/SAR) dispersed within more extended regions in other "boundary" domains, is discussed in relation to previously proposed models for insulation.

Animals↗

PubMed2004

Perioperative thermal insulation: minimal clinically important differences?

BACKGROUND: Reduction of heat losses from the skin by thermal insulation is used to avoid perioperative hypothermia. However, there is little information about the physical properties of various insulating materials used in the operating room. METHODS: The following insulation materials were tested using a validated manikin: cotton surgical drape tested in two and four layers; Allegiance drape; 3M Steri-Drape; metallized plastic sheet; Thermadrape Barkey thermcare 1 tested in one and two layers; hospital duvet tested in one and two layers. Heat loss from the surface of the manikin can be described as: Q(*);= h.DeltaT.A where Q(*); is heat flux, h is the heat exchange coefficient, DeltaT is the temperature gradient between the environment and surface and A is the area covered. The heat flux per unit area (Q(*); A(-1)) and surface temperature were measured with nine calibrated heat-flux transducers. The environmental temperature was measured using a thermoanemometer. DeltaT was varied and h was determined by linear regression analysis as the slope of DeltaT vs Qdot; A(-1). The reciprocal of h defines the insulation. RESULTS: The insulation value of air was 0.61 Clo. The insulation values of the materials varied between 0.17 Clo (two layers of cotton surgical drapes) to 2.79 Clo (two layers of hospital duvet). CONCLUSIONS: There are relevant differences between various insulating materials. The best commercially available material designed for use in the operating room (Barkey thermcare 1) can reduce heat loss from the covered area by 45% when used in two layers. Given the range of insulating materials available for outdoor activities, significant improvement in insulation of patients in the operating room is both possible and desirable.

Air↗

PubMed1991

Resultant clothing insulation during exercise in the cold.

Predictions of required clothing during work in the cold is often based on data obtained from thermal manikin measurements. These clothing basic insulation values are altered during exercise. In a series of experiments at our laboratory, resultant clothing insulation has been studied. In a cold climatic chamber subjects walked on a treadmill at various exercise intensities dressed in cold protective clothing at different levels of insulation. Thermal insulation of the clothing ensembles was measured with a thermal manikin under standardized conditions. Compared to the standard manikin insulation values, the clothing insulation values measured (resultant insulation) at the human experiments were always lower. Reductions of insulation ranged from 12 to 54% depending on such factors as work intensity, sweating amount, clothing type etc. Predictions of cold protection of clothing at actual use must take into account factors influencing clothing insulation to avoid over-estimation of the cold protective properties of clothing.

Clothing↗

PubMed1985

The effect of leakage on the insulation provided by immersion-protection clothing.

The effect of controlled, incremental water leakage on the thermal insulation provided by three immersion-protection assemblies has been measured using a thermal manikin. The results show an average loss of 30% of the initial insulation for a leak of 500 g, 40% for a leak of 1000 g, and nearly 60% for a leak of 3000 g. The assemblies differed only in the thermal insulation layer which consisted of: A. A single thickness of wool (initial immersed insulation 0.63 clo) B. A double thickness of wool (initial immersed insulation 0.79 clo) and C. A layer of polypropylene batting between nylon covers (initial immersed insulation 0.76 clo). Differences between the assemblies in loss of insulation with leakage were small and no evidence was found to support claims that the insulating properties of polypropylene are more resistant to wetting than those of wool. It is considered that the substantial loss of insulation even with small leaks makes it essential that tests of the water-excluding performance of immersion suits are undertaken in realistic conditions rather than in calm water.

Body Temperature Regulation↗

PubMed2006

Identification, genome mapping, and CTCF binding of potential insulators within the FXYD5-COX7A1 locus of human chromosome 19q13.12.

Identification of insulators is one of the most difficult problems in functional mapping of genomes. For this reason, up to now only a few insulators have been described. In this article we suggest an approach that allows direct isolation of insulators by a simple positive-negative selection based on blocking enhancer effects by insulators. The approach allows selection of fragments capable of blocking enhancers from mixtures of genomic fragments prepared from up to 1-Mb genomic regions. Using this approach, a 1-Mb human genome locus was analyzed and eight potential insulators were selected. Five of the eight sequences were positioned in intergenic regions and two were within introns. The genes of the alpha-polypeptide H+/K+ exchanging ATPase (ATP4A) and amyloid beta (A4) precursor-like protein 1 (APLP1) within the locus studied were found to be flanked by insulators on both sides. Both genes are characterized by distinct tissue-specific expression that differs from the tissue specificity of the surrounding genes. The data obtained are consistent with the conception that insulators subdivide genomic DNA into loop domains that comprise genes characterized by similar expression profiles. Using chromatin immunoprecipitation assay, we demonstrated also that at least six of the putative insulators revealed in this work could bind the CTCF transcription factor in vivo. We believe that the proposed approach could be a useful instrument for functional analysis of genomes.

Animals↗

PubMed2006

A DNA insulator prevents repression of a targeted X-linked transgene but not its random or imprinted X inactivation.

Some genes on the inactive X chromosome escape silencing. One possible escape mechanism is that heterochromatization during X inactivation can be blocked by boundary elements. DNA insulators are candidates for blocking because they shield genes from influences of their chromosomal environment. To test whether DNA insulators can act as boundaries on the X chromosome, we inserted into the mouse X-linked Hprt locus a GFP transgene flanked with zero, one, or two copies of a prototypic vertebrate insulator from the chicken beta-globin locus, chicken hypersensitive site 4, which contains CCCTC binding factor binding sites. On the active X chromosome the insulators blocked repression of the transgene, which commences during early development and persists in adults, in a copy number-dependent manner. CpG methylation of the transgene correlated inversely with expression, but the insulators on the active X chromosome were not methylated. On the inactive X chromosome, insulators did not block random or imprinted X inactivation of the transgene, and both the insulator and transgene were almost completely methylated. Thus, the chicken hypersensitive site 4 DNA insulator is sufficient to protect an X-linked gene from repression during development but not from X inactivation.

Animals↗

PubMed2000

Evaluation of heterologous insulator function with regard to chromosomal position effect in the mouse blastocyst and fetus.

Insulators are located at the boundaries of differentially regulated genes and delimit their interactions by establishing independent chromatin structures. Recently, an insulator sequence has been found in the 5'-flanking region of arylsulfatase (ARS) gene from sea urchin. To investigate functional conservation of this ARS insulator in mice, we performed blastocyst assays to evaluate the effect of this insulator on the chromosomal position effect, quantitatively. We constructed transgenes that have a luciferase gene under the control of the CMV-IE enhancer and the human elongation factor 1 alpha promoter in the presence or absence of the ARS insulator in both flanking regions. These transgenes were microinjected into 1-cell mouse embryos and luciferase activity was measured at the blastocyst stage. We found that the presence of ARS insulator sequence doubled the number of luciferase-expressing blastocysts, and that the proportion of the blastocysts with high-level expression (> or = 1 x 10(4) relative light units (RLU)) was increased more than tenfold. In the case of transgenic fetuses, however, the presence of ARS insulator did not seem to improve transgene expression. These results suggest that the sea urchin ARS insulator confers position-independent expression driven by the human elongation factor 1 alpha promoter, at least in the blastocyst stage of the mouse.

Animals↗