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Expression of Reg gene in the Syrian golden hamster pancreatic islet regeneration model.

We have reported previously that cellophane wrapping of the hamster pancreas is a stimulus that leads to the induction of duct epithelial cell proliferation, followed by endocrine cell differentiation and new islet formation. Reg is a candidate gene that has been reported to be expressed in regenerating pancreatic islets, suggesting a role in islet growth. We examined Reg gene expression in the cellophane-wrap model by isolating total RNA from hamster pancreata at various times after wrapping. Northern blot analysis using a rat cDNA Reg probe showed no expression of Reg in control non-wrapped hamster pancreas, whereas a strong signal was detected in control wrapped rat pancreas. Using reverse transcription of RNA followed by polymerase chain reaction (PCR) we amplified, isolated and sequenced a 194 base pair product which showed homology to rat Reg in both control and wrapped hamster pancreas. When the PCR product was used as a probe for Northern blot analysis, no signal was detected in control non-wrapped pancreata. In contrast, a strong signal was detected 1 and 2 days after wrapping, which then returned to basal between 4 and 6 days after wrapping. A similar temporal pattern was observed using in situ hybridization to localize the Reg gene. One- and 2-day wrapped but not control pancreas expressed Reg in acinar cells, but not in islets.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Prevalence of intestinal parasite infections on a national scale among primary schoolchildren in Laos.

In order to investigate the epidemiological situation of intestinal parasite infections in Laos, parasitological surveys were carried out on a national scale including 17 provinces and the Vientiane Municipality. A total of 29,846 stool specimens were collected from primary schoolchildren from May 2000 to June 2002 and examined once with the cellophane thick smear technique. The cumulative egg positive rate for intestinal helminths was 61.9%. By species, the rate for Ascaris lumbricoides was 34.9%, hookworm 19.1%, Trichuris trichiura 25.8%, Opisthorchis viverrini 10.9%, Taenia spp. 0.6% and Hymenolepis spp. 0.2%. The northern mountainous regions such as Phongsaly, Huaphan or Saysomboune Province showed a higher prevalence (over 70%) of soil- transmitted helminths. The regions along the Mekong River such as Khammuane, Saravane or Savannakhet Province showed a higher prevalence (over 20%) of fish-borne parasites. On the other hand, Schistosoma mansoni eggs were detected in 1.7% of schoolchildren only in Champassak Province, a previously endemic area. The highest prevalence was noted in Phongsaly Province (96.0%) and the lowest in Bolikhamxay Province (27.5%). An additional small-scale survey by cellophane anal swab detected Enterobius vermicularis eggs in 35.7% of 451 schoolchildren aged 6-8 years in Khammuane, Vientiane, Champassak Province and the Vientiane Municipality. Meanwhile, the mean blood haemoglobin level of hookworm-infected children was not lower than that of children not infected with hookworm, suggesting that nutritional factors are more important than parasite infection per se. Nevertheless, the above results indicate that a nationwide parasite control project is necessary to reduce possible morbidity due to parasitic diseases in the country.

Child↗

Development of experimental cancer in the head of the pancreas by surgical induction of tissue injury.

The majority of carcinomas of the pancreas in humans are of ductal origin and are located in the head of the gland. These clinical characteristics however, are not affected in traditional animal models of the disease. Partial pancreatic duct obstruction with ductal epithelial hyperplasia was produced in the Syrian golden hamster by wrapping the head of the pancreas with cellophane tape. Wrapped and unwrapped animals were then exposed to an exogenous carcinogen (N-nitrosobis[2-oxopropyl]amine). Assay of serum ribonuclease activity was used as a marker of disease. Invasive lesions developed in both groups of animals. Fifty percent of the tumors in the Group II (cellophane wrap and N-nitrosobis [2-oxopropyl] amine) hamsters were located in the head of the gland and were of ductal origin. All tumors in animals receiving N-nitrosobis [2-oxopropyl]amine alone (Group I) occurred peripherally and were derived from ductular or acinar tissue. Elevation of serum ribonuclease activity was noted early in the course of carcinogenesis, thereafter returning to normal, which may explain the clinical controversy regarding this marker. This new model should enhance our knowledge of the interrelationships between etiologic factors, precursor lesions, and pancreatic cancer.

Animals↗

Bacteriophage Mu: a transposing replicon.

Mu DNA replication has been carried out in vitro on cellophane discs in the presence of dBUTP. If the DNA is sheared to 80 kb pieces, the Mu replicas band anomalously in CsCl gradients between hybrid and light DNA density positions. The intermediate density DNA comprises semiconservatively replicated Mu sequences, flanked by unreplicated light DNA. This and previous data are consistent with replication occurring within Mu boundaries. Both the synthesis of Mu sequences and the intermediate density DNA are abolished by protein synthesis inhibition in vivo just prior to lysis on cellophane discs. These observations indicate that at least some steps in bona fide Mu transposition-replication are being observed in vitro.

Bacteriophage mu↗

Complement, granulocytes, and shock lung.

Intravascular leukostasis in the pulmonary microvasculature is a cardinal early histologic finding in patients with shock lung. Identical leukostasis is also observed in patients undergoing extracorporeal hemodialysis with cellophane membrane dialyzers, and it has been documented that the accumulation of granulocyte plugs in the lung is mediated by complement activation triggered by dialyzer cellophane. The C5a-desarg so generated causes peripheral blood granulocytes to aggregate, and the aggregates so formed embolize to the lung, where they cause occlusion of the microvasculature and increased capillary leakage, manifested by interstitial and alveolar edema. In vitro studies suggest that this endothelial damage is mediated by hydrogen peroxide from the adherent granulocytes. Most importantly, a close correlation has been found between the presence of C5a-desarg in plasma and the subsequent onset of shock lung in patients after trauma, burns, and sepsis. As exemplified by hemodialysis leukopenia, C5a-desarg-mediated pulmonary leukostasis is a self-limiting process because of selective down-regulation of granulocyte receptors for C5a-desarg--a mechanism that primarily limits the lung damage associated with intravascular complement activation.

Animals↗

Enhancement of the release of azelaic acid through the synthetic membranes by inclusion complex formation with hydroxypropyl-beta-cyclodextrin.

The aim of this study was to investigate the release rates of azelaic acid and azelaic acid-hydroxypropyl-beta-cyclodextrin (HPbetaCD) inclusion complex through three types of synthetic membranes, namely cellophane, silicone and elastomer membranes. Solid inclusion complexes of azelaic acid-HPbetaCD at the molar ratio of 1:1 were prepared by coevaporation and freeze-drying methods, subsequently characterized by differential scanning calorimetry, X-ray diffractometry and dissolution studies. Solid inclusion complex obtained by coevaporation method which exhibited the inclusion of azelaic acid in the HPbetaCD cavity and gave the highest dissolution rate of azelaic acid was selected for the release study. Release studies of azelaic acid and this complex through the synthetic membranes were conducted using vertical Franz diffusion cells at 30 degrees C for 6 days. The release rates of azelaic acid through the synthetic membranes were enhanced by the formation of inclusion complex with HPbetaCD at the molar ratio of 1:1, with the increasing fluxes of about 41, 81 and 28 times of the uncomplexed system in cellophane, silicone and elastomer membranes, respectively. The result from this study can be applied for the development of azelaic acid for topical use.

2-Hydroxypropyl-beta-cyclodextrin↗

Development and evaluation of an artificial membrane for determination of drug availability.

Various artificial membranes (e.g. PAMPA) and cellular-based membranes (e.g. Caco-2) are used for screening during early stages of drug discovery. However, these methods are not well suited for evaluation of pharmaceutical formulations and the effects of various excipients on drug availability. When drug molecules permeate biological membranes they encounter two types of permeation resistance, a membrane resistance in the lipophilic membrane and diffusion resistance in the unstirred water layers adjacent to both surfaces of the lipophilic membrane. We have developed an artificial membrane that is cheap and simple to prepare. The unstirred water layer consists of a hydrated semi-permeable cellophane membrane with a molecular weight cutoff (MWCO) of 12,000-14,000 Da and a lipophilic membrane of pure n-octanol in a nitrocellulose matrix. In the diffusion cell the hydrated cellophane membrane (thickness 210-230 microm) is on the donor side and the lipophilic octanol membrane (thickness about 120 microm) on the receptor side. Permeation of ionizable lipophilic drug molecules was diffusion-controlled when the drug was unionized but lipophilic membrane controlled when the drug was ionized. Drug permeation patterns from cyclodextrin containing formulations through the membrane were similar to those previously observed for biological membranes such as hairless mouse skin and the eye cornea.

Animals↗

Plasma formation and shielding by three ophthalmic neodymium-YAG lasers.

Because microplasma formation may provide an energy-absorbing shield that protects the retina from inadvertent injury during neodymium-YAG laser surgery, we studied plasma formation and shielding by three ophthalmic neodymium-YAG lasers (the Q-switched American Medical Optics YAG-100, the mode-locked Meditec OPL-3, and the Q-switched LASAG Sirius Microruptor 2) with three model targets (physiologic saline, 1% sodium hyaluronate, and a cellophane membrane). Plasma formation produced a linear reduction in the transmission of light energy along the beam path in a qualitatively differences. Plasma shielding was similar in both saline and sodium hyaluronate; enhancement of the shield effect occurred only with the cellophane membrane target. Operating the lasers at energy levels far above threshold did not reduce total light transmission significantly. Clinical use should be guided by the principle of employing the minimum energy necessary to cut the target tissue.

Lasers↗

Prevalence and associations of epiretinal membranes. The Blue Mountains Eye Study, Australia.

PURPOSE: The purpose of the study is to determine the prevalence and associations of epiretinal membranes in a defined older Australian population and to assess their influence on visual acuity. METHODS: Three thousand six hundred fifty-four persons 49 years of age or older, representing 88% of permanent residents from an area west of Sydney, underwent a detailed eye examination, including stereo retinal photography. Epiretinal membranes were diagnosed clinically and from photographic grading. RESULTS: Signs of epiretinal membranes were found in 243 participants (7%; 95% confidence interval [CI], 6.1, 7.6), bilateral in 31%. The prevalence was 1.9% in persons younger than 60 years of age, 7.2% in persons 60 to 69 years of age, 11.6% in persons 70 to 79 years of age, and 9.3% in persons 80 years of age and older, with slightly higher rates in women. Two stages were identified: an early form without retinal folds, termed "cellophane macular reflex" present in 4.8%, and a later stage with retinal folds, termed "preretinal macular fibrosis" (PMF), found in 2.2% of the population. Preretinal macular fibrosis, but not cellophane macular reflex, had a small, significant effect on visual acuity. Preretinal macular fibrosis was significantly associated with diabetes, after age-gender adjustment, in subjects without signs of diabetic retinopathy (odds ratio, 3.2; 95% CI, 1.4, 7.2). Preretinal macular fibrosis also was associated with increased fasting plasma glucose (odds ratio, 1.2; 95% CI, 1.1, 1.3). Epiretinal membranes were found in 16.8% of persons who had undergone cataract surgery in one or both eyes (including PMF in 3.7%), in 16.1% of retinal vein occlusion cases (PMF in 12.5%), both significantly higher rates than in subjects without these conditions (P < 0.0001), and in 11% of persons with diabetic retinopathy (PMF in 3.6%), not significantly higher (P = 0.17). CONCLUSIONS: This study has documented the frequency and mild effect on vision of epiretinal membranes in an older population. Diabetes was associated significantly with idiopathic cases, whereas well-known associations with past cataract surgery and retinal disease were confirmed.

Adult↗

Is scleral fixation a safe procedure for intraocular lens implantation?

PURPOSE: No consensus currently exists on the optimal method for intraocular (IOL) implantation without capsular support. We evaluated the outcome and angiographic findings of eyes that underwent the implantation of scleral fixated IOLs. METHODS: Iris and retinal fluorescein angiography were performed in 13 eyes that had received posterior chamber IOL implantation with scleral fixation. Follow-up examinations also assessed visual acuity (VA), intraocular pressure (IOP), IOL decentration and complications related to the procedure. RESULTS: Mean visual acuity was 0.29 preoperatively and 0.71 postoperatively after a mean follow-up of 14.2 months. A best corrected visual acuity of 0.5 or better was obtained in 12 eyes. Iris fluorescein angiography did not show major vascular abnormalities. Retinal angiography showed 5 cases of macular edema. In 6 eyes light-induced retinal lesions occurred. Cellophane maculopathy was disclosed in 4 eyes. Macular edema was associated with photic injury in 4 cases and with cellophane maculopathy in 2 cases. Mean postoperative visual acuity was 0.6 in eyes with macular edema and 0.88 in eyes without (SD 0.18; range 0.5-1.0). Four of 5 eyes with macular edema had a postoperative visual acuity of 0.5 or better. There was no evidence of persistent IOP elevation or IOL decentration. No serious complications were recorded during surgery. CONCLUSIONS: Transscleral fixation of posterior chamber IOLs provides adequate visual acuity in most patients. Macular edema was frequently associated with the procedure. Although this complication was a cause of low visual recovery after implantation, the majority of eyes with macular edema achieved a visual acuity of 0.5 or better. Light-induced retinal injury was a permanent complication.

Aged↗

[Is removal of internal limiting membrane always necessary during surgery for refractory diffuse diabetic macular edema without evident epimacular proliferation?].

PURPOSE: The aim of this study was to evaluate the effect of the internal limiting membrane (ILM) peeling in eyes with diabetic macular edema (DME) but without evident epimacular proliferation or cellophane maculopathy that were unresponsive to laser photocoagulation. Secondly we wanted to determine whether ILM peeling is always essential in DME surgery and whether it improves the functional outcome. PATIENTS AND METHODS: In a prospective controlled study, ten patients with similar degrees and duration of DME in both eyes were followed-up for more than 6 months after bilateral vitrectomy with and without ILM peeling. Trypan blue 0.2 % was used to stain the ILM during surgery. We evaluated the anatomic outcome as detected by bimicroscopic evaluation and optical coherence tomography (OCT) and the visual outcome. Intraoperatively and postoperatively any complications occurring were documented. RESULTS: Baseline BCVA and foveal thickness ranged, respectively, from 20/50 to 20/400 (mean 20/110) and 430 to 840 microm (mean 618) in eyes that underwent ILM peeling, 20/40 to 20/400 (mean 20/120) and 390 to 910 microm (mean 623 microm) in eyes without ILM peeling. There were no significant differences between the both groups in baseline BCVA (P = 0.4691, Wilcoxon rank sum test) or foveal thickness (P = 0.8204, Wilcoxon rank sum test). At six-months follow-up, mean BCVA improved significantly in both groups, from 20/110 to 20/60 (P = 0.0427, Wilcoxon signed rank test) in eyes that underwent ILM peeling and from 20/120 to 20/80 (P = 0.0482, Wilcoxon signed rank test) in eyes without ILM peeling. Mean foveal thickness decreased significantly from 618 to 265 (P = 0.0050) in eyes with ILM peeling and from 623 to 311 (P = 0.0050) in eyes without ILM peeling. Visual acuity improved by two or more lines in five eyes (50 %) of each group. There were no significant differences in the improvement of BCVA and decreasing of foveal thickness between the both groups (Wilcoxon rank sum test, P = 0.9083, P = 0.2720, respectively). Cyst rupture with formation of macular hole was documented in one eye after ILM peeling. CONCLUSIONS: Vitrectomy with or without ILM peeling may improve BCVA and decrease foveal thickness. ILM peeling was not found to enhance the improvement of VA postoperatively. A larger study is required to determine whether ILM peeling is essential in surgery for DME without epimacular proliferation or cellophane maculopathy.

Diabetic Retinopathy↗

DNA intermediates at the Escherichia coli replication fork: effect of dUTP.

We have directly tested the hypothesis that elevated levels of dUTP cause the formation of small DNA fragments at the replication fork of Escherichia coli. Addition of increasing levels of dUTP to lysates on cellophane discs results in an increasing number of strand scissions in the newly replicated DNA. Lysates of strains defective in dUTPase produce many more scissions at the same level of dUTP. The size distribution of Okazaki pieces obtained in vivo can be reconstituted in vitro on cellophane discs if appropriate levels of dUTP are present. Although uracil excision leads to the apparent production of Okazaki pieces from both daughter strands, DNA synthesis is actually asymmetric under these conditions. De novo chain initiation events occur on only one strand. It is suggested that asymmetry of synthesis in vivo may be masked by uracil excision and other postreplication processing mechanisms.

DNA Polymerase I↗

An epidemiological study of Fasciola hepatica in The Netherlands.

Transmission of F. hepatica under natural conditions was analysed in a three year programme. The variables used were the indirect haemagglutination (IHA) technique, worm establishment in tracer lambs and the population dynamics, infection rate and shedding pattern of Lymnaea truncatula. It is concluded that fluke eggs, infected snails and metacercariae on herbage can survive the winter in the Netherlands. Metacercarial availability was positively correlated to the amount of rainfall in the grazing period. The role developed eggs that survive the winter is important, because this results in earlier infections in the herd. The use of the serological diagnosis method IHA is important to detect F. hepatica infection in an early stage. Use of cellophane paper on floats is a useful method for determining the shedding pattern of cercariae from L. truncatula. It is concluded that collection of metacercariae on cellophane floats, inventarization of L. truncatula and its infection level are useful tools for the prediction of liverfluke infections.

Animals↗

Alterations of the glomerular beta-adrenoceptor-linked adenylate cyclase system in perinephritis hypertension.

The present study was undertaken to investigate beta-adrenoceptor-adenylate cyclase coupling in a myocardial ventricular membrane preparation and in isolated renal glomeruli of cellophane perinephritis hypertensive rats. Adenylate cyclase activity and [125I]iodocyanopindolol binding were differentially affected in these preparations. In isolated glomeruli of hypertensive rats a reduced intrinsic activity of isoproterenol was associated with an apparent loss of the guanine nucleotide-sensitive high-affinity state of the beta-adrenoceptors, whereas their absolute number was unchanged when compared with the normotensive control rats. On the other hand, in the sarcolemmal preparation of hypertensive rats adenylate cyclase activity, the relative amount of high-affinity states, and the density of beta-adrenoceptors were not different from the respective values in normotensive controls. Experiments performed on isolated glomeruli of rats with unilateral cellophane perinephritis that developed only moderate hypertension provide evidence that the apparent loss of the high-affinity state is a consequence of hypertension, since no difference was observed in glomeruli from the wrapped, as compared with the intact kidney.

Adenylyl Cyclases↗

Auxiliary method for clonal identification of Staphylococcus aureus by protein band pattern of released proteins on SDS-polyacrylamide gel.

A supportive method for clonal identification of Staphylococcus aureus strains was devised. Culture supernatant obtained by cellophane surface culture was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) without performing any concentration procedure prior to electrophoresis. The combined use of cellophane surface culture and SDS-PAGE was convenient for determining whether the strains belonged to the same clone or not when conducted in conjunction with other tests for bacteriological characterization.

Bacterial Proteins↗

Differential effect of renal wrap hypertension on aortic smooth muscle polyploidy in the rat and rabbit.

1. The incidence of aortic smooth muscle cell polyploidy was investigated in rabbits and rats with renal wrap induced (cellophane perinephritic) chronic hypertension. 2. Bilateral renal cellophane wrapping was performed in young adult animals. Blood pressure was measured intra-arterially in the rabbits twice during the experimental period and tail-cuff blood pressure measured twice weekly in the rats. At 8 weeks post-surgery the incidence of aortic smooth muscle cell polyploidy was determined in enzymatically isolated cells by flow cytometric DNA analysis. 3. Systolic blood pressure was significantly increased in the bilateral renal wrapped rabbits and rats compared to the shams, such that at 8 weeks post-surgery, systolic blood pressure was 139 +/- 2 mmHg and 84 +/- 2 mmHg, respectively, in the rabbits and 188 +/- 6 mmHg and 155 +/- 4 mmHg, respectively, in the rats. 4. The incidence of polyploid smooth muscle cells was significantly higher in the hypertensive renal wrapped rat compared to the sham (20.9 +/- 1.5% and 8.1 +/- 0.5%, respectively). However, the incidence of polyploid cells was low in the rabbit aortae with no significant difference in the incidence of aortic smooth muscle polyploidy in the hypertensive rabbit compared to the sham (2.6 +/- 0.6% and 2.7 +/- 0.6%, respectively). 5. This study demonstrates a species difference in the induction of polyploidy during the same model of experimental hypertension in aortic smooth muscle derived from the rabbit and rat.

Animals↗

Chronic hydronephrosis: renographic drainage patterns and renal morphology in an animal model.

OBJECTIVE: To develop an animal model of chronic progressive hydronephrosis; to establish patterns of diuretic renography during development of the condition; and to relate renographic abnormality to morphological change in the renal parenchyma. MATERIALS AND METHODS: The cellophane sclerosis technique was used to produce a hydronephrosis in the left kidney of six rabbits. Post-operatively MAG3 diuretic renography was performed at weekly intervals. When a steady renographic trend was noted the rabbits were killed and the kidneys examined histologically. RESULTS: Four rabbits had persistent type II renographic curves on the left for 12 weeks. The measured split renal function fell steadily to < 20% in this time. Morphologically there was hydronephrosis with marked structural changes in the parenchyma seen on histological examination. Two rabbits had stable type IIIa renographic drainage patterns on the left with preserved renal function. Morphologically there was hydronephrosis but histologically the kidneys were normal. There were no renographic or structural changes in a control rabbit after a sham laparotomy. CONCLUSIONS: The cellophane sclerosis technique can be used to create a reproducible chronic progressive hydronephrosis. In this animal model hydronephrosis and a persistent type II curve are associated with renographic evidence of deteriorating renal function and disruption of renal architecture histologically. Hydronephrosis and a type IIIa curve are associated with preservation of normal renal function and histological structure.

Animals↗

Differential expression of fungal genes at preinfection and mycorrhiza establishment between Terfezia boudieri isolates and Cistus incanus hairy root clones.

Changes in gene expression by isolates of Terfezia boudieri during mycorrhization with Cistus incanus hairy roots were followed. Four fungus-hairy root clone combinations were cultivated under two sets of conditions, in which the root and the fungus were separated by a cellophane sheet or were allowed physical contact. One of the combinations produced endomycorrhizas, the other three solely ectomycorrhizas. Fragments isolated by cDNA-AFLP analysis from cellophane-separated cultures (preinfection) were used to identify differentially expressed genes by reverse Northern analysis. Genes showing no homology to known sequences constituted the largest group under both growth conditions. Some fungal genes were expressed transiently, while others exhibited altered expression patterns as conditions changed from individually growing through the preinfection stage to mycorrhizas. Genes expressed exclusively under combinations allowing either ectomycorrhiza or endomycorrhiza under a particular condition were detected. Our results point, for the first time, to some of the genes that might be involved in determining the type of association that will be formed: ecto- or endomycorrhiza.

Ascomycota↗