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Biomedical subjects

E Winkler

Publications and source records attributed to E Winkler.

At least 19 recordsLinked to original sources

Sexual and vegetative reproduction of Hieracium pilosella L. under competition and disturbance: a grid-based simulation model.

We used a spatially explicit simulation model to examine the relative importance of vegetative and sexual reproduction in Hieracium pilosella L. Based on an understanding of the complex life cycle of this species and on data from in situ population dynamics in a calcareous grassland in NW Switzerland, we simulated growth and the relative contribution of clonal reproduction by stolons and reproduction by seeds across a gradient of increasing soil fertility. Competition by a clonal grass resulted in nearly complete exclusion of H. pilosella from the more fertile part of the simulation plot. Under low soil fertility, when grass could not survive, H. pilosella largely persisted by vegetatively produced rosettes. This pattern of a sharp separation of both species was shifted slightly in favour of H. pilosella by introducing random disturbances. Only by adding: (1) long-distance seed dispersal, and (2) facilitation of seedling establishment in the vicinity of grass tussocks in vegetation gaps was a more realistic representation of field observations realised, with rosettes of H. pilosella grown from seeds occasionally distributed within dense grass vegetation. Phenotypic plasticity of stolon length was a decisive factor for the maintenance of H. pilosella populations. We conclude that a mixed strategy of clonal growth and reproduction by seeds in H. pilosella is necessary to maintain populations of this species in the presence of high interspecific competition and a shortage of open space.

Algorithms↗

UCP3 expressed in yeast is primarily localized in extramitochondrial particles.

Previously it was concluded (1) that, differently from UCP1, on expression in Saccharomyces cerevisiae, UCP3, and UCP3 short (UCP3s) are in a deranged state, allowing for unregulated uncoupling. Here we show that the bulk of UCP3 and UCP3s is in extramitochondrial aggregates whether expressed with high or medium expression vectors. The evidence is based on the insolubility of most UCP3 and UCP3s in nonionic detergents such as Triton X100, in contrast to UCP1. Using very high expression vector, macroscopic evidence for extramitochondrial UCP3 containing particles is a viscous white sediment surrounding the mitochondrial fraction which contains UCP3 as inclusion body type aggregate. Together with the previous data it is concluded that uncoupling due to small amounts of incorporated, deranged, and nucleotide insensitive UCP3 prevents incorporation of the bulk of UCP3 into mitochondria. This finding also provides a simple and stringent assay for the state of heterologously expressed in mitochondrial membrane proteins.

Carrier Proteins↗

Uncoupling proteins 2 and 3 are highly active H(+) transporters and highly nucleotide sensitive when activated by coenzyme Q (ubiquinone).

Based on the discovery of coenzyme Q (CoQ) as an obligatory cofactor for H(+) transport by uncoupling protein 1 (UCP1) [Echtay, K. S., Winkler, E. & Klingenberg, M. (2000) Nature (London) 408, 609-613] we show here that UCP2 and UCP3 are also highly active H(+) transporters and require CoQ and fatty acid for H(+) transport, which is inhibited by low concentrations of nucleotides. CoQ is proposed to facilitate injection of H(+) from fatty acid into UCP. Human UCP2 and 3 expressed in Escherichia coli inclusion bodies are solubilized, and by exchange of sarcosyl against digitonin, nucleotide binding as measured with 2'-O-[5-(dimethylamino)naphthalene-1-sulfonyl]-GTP can be restored. After reconstitution into vesicles, Cl(-) but no H(+) are transported. The addition of CoQ initiates H(+) transport in conjunction with fatty acids. This increase is fully sensitive to nucleotides. The rates are as high as with reconstituted UCP1 from mitochondria. Maximum activity is at a molar ratio of 1:300 of CoQ:phospholipid. In UCP2 as in UCP1, ATP is a stronger inhibitor than ADP, but in UCP3 ADP inhibits more strongly than ATP. Thus UCP2 and UCP3 are regulated differently by nucleotides, in line with their different physiological contexts. These results confirm the regulation of UCP2 and UCP3 by the same factors CoQ, fatty acids, and nucleotides as UCP1. They supersede reports that UCP2 and UCP3 may not be H(+) transporters.

Animals↗

Treatment of rhinophyma with Er:YAG laser.

BACKGROUND AND OBJECTIVE: Rhinophyma is a benign, disfiguring disorder of the nose which presents the end stage of acne rosacea, and can also occasionally result in nasal airway obstruction. We describe the first series of patients treated with Erbium laser. STUDY DESIGN/MATERIALS AND METHODS: Severe-to-moderate rhinophyma in six patients were treated at our institution between 1995 and 1996, using the Erbium:YAG (Erb:YAG) laser. RESULTS: All patients achieved marked cosmetic improvement, with no complications. Post-operative healing time was seven to fourteen days-significantly shorter than similar other modalities. CONCLUSIONS: The Erbium laser provides very accurate tissue ablation and allows the sculpturing of the hypertrophied areas, offering good cosmetic results, with a very short healing period as shown in our study.

Aged↗

Endoproteolysis of the ER stress transducer ATF6 in the presence of functionally inactive presenilins.

Presenilin (PS) proteins facilitate endoproteolysis of selected type I transmembrane proteins such as the Alzheimer's disease (AD) associated beta-Amyloid precursor protein (beta APP) and Notch. beta APP is cleaved within its transmembrane domain by an aspartyl protease activity termed gamma-secretase, which may be identical with PS1 and PS2. Notch also undergoes a PS-dependent intramembraneous proteolysis. A similar gamma-secretase-like cleavage may also occur with IRE1 and ATF6, two signaling molecules of the unfolded protein response (UPR) that may require PSs for their activation. Here, we have analyzed whether ATF6 cleavage requires a PS-dependent gamma-secretase activity and whether inhibition of gamma-secretase activity would affect the UPR. Endoproteolysis of ATF6 was observed in the presence of the highly potent gamma-secretase inhibitor L-685,458. ATF6 processing also occurred in the presence of functionally inactive dominant negative mutants of PS1 (PS1 D385N) and PS2 (PS2 D366A) that do not support endoproteolysis of beta APP and Notch. Our results therefore demonstrate that ATF6 is not a substrate for PS mediated gamma-secretase-like endoproteolysis. This finding indicates that gamma-secretase inhibitors, which are currently developed as therapeutic agents to lower the A beta burden in brains of AD patients, do not interfere with the UPR response.

Activating Transcription Factor 6↗

Uncoupling protein, H+ transport and regulation.

The biochemical functions of uncoupling proteins (UCPs) are discussed with the view of UCP1 as a paradigm. In contrast with UCP1, the heterologous expression of UCP3 in yeast is found to result primarily in extra-mitochondrial deposits and thus is unsuitable for studying UCP3 function. On expression in Escherichia coli inclusion bodies, UCPs extracted and incorporated into vesicles showed no H(+) transport, only Cl(-) transport. Only after addition of coenzyme Q was fully nucleotide-sensitive high-H(+) transport reconstituted, with UCP1 as well as with UCP2 and UCP3. The newly discovered cofactor role of coenzyme Q in H(+) transport is proposed to imply co-operation with fatty acids for the injection of H(+) into the UCP channel.

Adenosine Triphosphate↗

Total lower lip reconstruction with innervated muscle-bearing flaps: a modification of the Webster flap.

BACKGROUND: Mohs surgery and other surgical techniques are used for the removal of squamous cell carcinoma of the lower lip and may leave a large defect in the vermilion and underlying tissue. When nearly the entire lower lip is excised, reconstruction of this defect is a challenge. Repair requires the matching of vermilion color, maintenance of oral sphincter function and mouth opening size, and retention of sensation. Several techniques have been suggested. We present a modification of the Webster flap for total lower lip reconstruction using innervated muscle-bearing flaps. OBJECTIVE: To present a surgical technique for the reconstruction of total lower lip defects after excision of squamous cell carcinoma. METHODS: Innervated muscle-bearing flaps are used and demonstrated in one case. The surgical technique is discussed in detail. RESULTS: The reconstructive results were excellent. There were no postoperative complications. CONCLUSION: The use of innervated muscle-bearing flaps is a useful and effective option for the reconstruction of total lower lip defects.

Carcinoma, Squamous Cell↗

The versatility of the nasolabial flap enhanced by the delay procedure.

BACKGROUND: The nasolabial flap is a versatile and effective option for the closure of nasal defects of the cheek and nasal sidewall following Mohs surgery. However, both extirpation of a tumor in the region of the base of the flap or previous use of the flap often destroy the proximal axial blood supply to the flap, excluding its immediate utilization. We describe a different use of the nasolabial flap and a technique for preserving its capacity in the case of reutilization OBJECTIVE: To describe the versatility of the nasolabial flap and a delay procedure that enables its exploitation despite prior disruption of the proximal blood supply. METHODS: Three different uses of the same nasolabial flap are demonstrated in one patient. The surgical techniques are discussed in detail. RESULTS: The reconstructive results were excellent. There were no postoperative complications. CONCLUSION: The nasolabial flap is a versatile and effective option for the closure of nasal defects of the cheek and nasal sidewall. When the nasolabial flap has been used before, or its blood supply compromised, the delay procedure can reestablish its applicability.

Basal Cell Carcinoma↗

Snakebite poisoning in children--a call for unified clinical guidelines.

In Israel, there are no uniform guidelines for the treatment policy of children snake-bitten by the Vipera palaestinae, the most abundant venomous snake in the country. We conducted a retrospective study aiming to compare treatment policies in two different medical centers. We found significant differences regarding admission and steroid administration criteria. Although the differences between the centers regarding anti-venom administration did not reach statistical significance, there were substantial differences. Neither of the centers had a well-established policy for the treatment of snake envenomation in children. In the era of cost containment, a policy of routine admission of children to the PICU service following V. palaestinae envenomation is unjustified, especially since the introduction of a specific monovalent anti-venom into the therapeutic armamentarium.

Adolescent↗

The use of magnetic resonance imaging in the differential diagnosis between starch and fecal peritonitis.

Granulomatous peritonitis, caused by the starch from the surgeons' gloves, is a hypersensitivity reaction that can complicate abdominal surgery and mimic other causes of peritonitis. The diagnosis of this entity is difficult to make, and relies on a high index of suspicion. We suggest the use of magnetic resonance imaging to facilitate the diagnosis of this condition, based on an experimental animal model. 84 rats were subjected to laparotomy, and the abdominal cavity was exposed to either saline solution, talc solution, starch solution or fecal material by creating a cecal perforation. TI-weighted magnetic resonance images, with and without gadolinum enhancement, were taken after 3, 5 and 10 days. The animals were then sacrificed and the abdominal contents were evaluated both macroscopically and microscopically. Both talc and starch caused gross adhesions involving the entire abdominal viscera, and microscopy revealed signs of inflammation and fibrosis. Starch induced reactive granulomas. The adhesions in the cases of fecal peritonitis were confined to the area of the perforation. MR images of the starch peritonitis group was remarkable for a diffuse pathological process with enhancement of the omentum and the peritoneum after gadolinum injection. The MR readings of the fecal peritonitis group showed a localized process with no diffuse enhancement of the peritoneum. The striking differences between the magnetic resonance images of starch and fecal peritonitis in rats suggest that this modality is both sensitive and specific in diagnosing starch peritonitis in a rat model. Early non-invasive diagnosis of these separate entities would ease the establishment of the appropriate treatment. We are currently investigating the use of MRI imaging in suspected starch peritonitis in humans.

Animals↗

Coenzyme Q is an obligatory cofactor for uncoupling protein function.

Uncoupling proteins (UCPs) are thought to be intricately controlled uncouplers that are responsible for the futile dissipation of mitochondrial chemiosmotic gradients, producing heat rather than ATP. They occur in many animal and plant cells and form a subfamily of the mitochondrial carrier family. Physiological uncoupling of oxidative phosphorylation must be strongly regulated to avoid deterioration of the energy supply and cell death, which is caused by toxic uncouplers. However, an H+ transporting uncoupling function is well established only for UCP1 from brown adipose tissue, and the regulation of UCP1 by fatty acids, nucleotides and pH remains controversial. The failure of UCP1 expressed in Escherichia coli inclusion bodies to carry out fatty-acid-dependent H+ transport activity inclusion bodies made us seek a native UCP cofactor. Here we report the identification of coenzyme Q (ubiquinone) as such a cofactor. On addition of CoQ10 to reconstituted UCP1 from inclusion bodies, fatty-acid-dependent H+ transport reached the same rate as with native UCP1. The H+ transport was highly sensitive to purine nucleotides, and activated only by oxidized but not reduced CoQ. H+ transport of native UCP1 correlated with the endogenous CoQ content.

Animals↗

The bulk of UCP3 expressed in yeast cells is incompetent for a nucleotide regulated H+ transport.

The impact of uncoupling protein (UCP) 1, UCP3 and UCP3s expressed in yeast on oxidative phosphorylation, membrane potential and H+ transport is determined. Intracellular ATP synthesis is inhibited by UCP3, much more than by UCP1, while similar levels of UCP3 and UCP1 exist in the mitochondrial fractions. Measurements of membrane potential and H+ efflux in isolated mitochondria show that, different from UCP1, with UCP3 and UCP3s there is a priori a preponderant uncoupling not inhibited by GDP. The results are interpreted to show that UCP3 and UCP3s in yeast mitochondria are in a deranged state causing uncontrolled uncoupling, which does not represent their physiological function.

Animals↗

Site-directed mutagenesis identifies residues in uncoupling protein (UCP1) involved in three different functions.

Using site-specific mutagenesis, we have constructed several mutants of uncoupling protein (UCP1) from brown adipose tissue to investigate the function of acidic side chains at positions 27, 167, 209, and 210 in H(+) and Cl(-) transport as well as in nucleotide binding. The H(+) transport activity was measured with mitochondria and with reconstituted vesicles. These mutant UCPs (D27N, D27E, E167Q, D209N, D210N, and D209N + D210N) are expressed at near wt levels in yeast. Their H(+) transport activity in mitochondria correlates well with the reconstituted protein except for D27N (intrahelical), which shows strong inhibition of H(+) transport in the reconstituted system and only 50% decrease of uncoupled respiration in mitochondria. In the double adjacent acidic residues (between helix 4 and helix 5), mutation of D210 and of D209 decreases H(+) transport 80% and only 20%, respectively. These mutants retain full Cl(-) transport activity. The results indicate that D210 participates in H(+) uptake at the cytosolic side and D27 in H(+) translocation through the membrane. Differently, E167Q has lost Cl(-) transport activity but retains the ability to transport H(+). The separate inactivation of H(+) and Cl(-) transport argues against the fatty acid anion transport mechanism of H(+) transport by UCP. The mutation of the double adjacent acidic residues (D209, D210) decreases pH dependency for only nucleoside triphosphate (NTP) but not diphosphate (NDP) binding. The results identify D209 and D210 in accordance with the previous model as those residues which control the location of H214 in the binding pocket, and thus contribute to the pH control of NTP but not of NDP binding.

Adipose Tissue, Brown↗

ESR/Alanine gamma-dosimetry in the 10-30 Gy range.

We report Alanine Dosimeter preparation, procedures for using the ESR/Dosimetry method, and the resulting calibration curve for gamma-irradiation in the range from 10-30 Gy. We use calibration curve to measure the irradiation dose in gamma-irradiation of human blood, as required in Blood Transfusion Therapy. The ESR/Alanine results are compared against those obtained using the thermoluminescent dosimetry (TLD) method.

Alanine↗

Two p16 (CDKN2A) germline mutations in 30 Israeli melanoma families.

Germline mutations in the p16 (CDKN2A) tumour suppressor gene have been linked to inherited predisposition to malignant melanoma (MM). Variable frequencies of p16 germline mutations were reported in different collections of melanoma families but it can be as high as 50%. Here we describe the results of p16 mutation screening in 30 melanoma kindreds in Israel. The entire coding region of the p16 gene, including exons 1, 2 and 3, flanking exon/intron junctions, and a portion of the 3' untranslated (UTR) region of the gene were examined by single-stranded conformation polymorphism (SSCP) analysis and direct sequencing. Two p16 germline mutations were identified: G101W, which has been previously observed in a number of melanoma kindreds, and G122V, a novel missense mutation. Thus, the frequency of mutations identified in this collection of Israeli families was 7%. Functional analysis indicated that the novel G122V variant retained some capacity to interact with cyclin dependent kinases (CDKs) in vitro, yet it was significantly impaired in its ability to cause a G1 cell cycle arrest in human diploid fibroblasts. This partial loss of function is consistent with the predicted impact of G122V substitution on the 3-dimensional structure of the p16 protein.

Adolescent↗

Photodynamic therapy of cutaneous lymphoma using 5-aminolevulinic acid topical application.

BACKGROUND: Photodynamic therapy (PDT) with topical application of 5-aminolevulinic acid (ALA) is a new and effective modality for treatment of superficial basal and squamous cell carcinomas. OBJECTIVE: We present the kinetics of ALA-induced protoporphyrin IX (PP) accumulation and the results of ALA PDT treatment on two patients with different stages (stage I and stage III) of mycosis fungoides (MF)-type cutaneous T-cell lymphoma (CTCL). METHODS: ALA-Decoderm cream was applied to the lesions for 16 hours. Spectrofluorescence measurements of PP accumulation were carried out before, during, and 1 hour after photoirradiation (580-720 nm) using the VersaLight system. RESULTS: Different patterns of PP fluorescence kinetics were observed in patients with early and advanced stages of the disease. During photoirradiation the intensity of fluorescence decreased depending on the lesion thickness. One hour after the photoirradiation procedure no PP fluorescence was observed in the stage I MF lesion, while in the thick stage III MF lesions, PP fluorescence reappeared; after an additional 10-15 minutes of irradiation PP fluorescence disappeared. Complete response with excellent cosmetic results was observed in the stage I lesion after a single irradiation with a light dose of 170 J/cm2; in five stage III lesions, complete response was achieved after fractionated irradiation with a total light dose of 380 J/cm2 (follow-up at 27 and 24 months, respectively). CONCLUSION: The results showed a high response of both stage I and stage III MF lesions to ALA PDT. This modality appears to be very effective and can be used successfully for MF treatment.

Abdomen↗

Triage for Leiurus quinquestriatus scorpion envenomation in children--is routine ICU hospitalization necessary?

(1) Leiurus quinquestriatus scorpion (LQS) envenomation is a common public health problem with a similar clinical presentation in the Middle East and worldwide: localized reactions occur in up to 97% of the victims. (2) LQS envenomation in children is potentially fatal since the severity of symptoms is weight-dependent. (3) A common policy is to hospitalize all children stung by the LQS-regardless of clinical severity-in the pediatric intensive care unit (PICU). (4) Seventeen of 18 children treated at two Israeli medical centers during an 8-year period developed mild to moderate clinical manifestations (antivenin was given in the one severe case; all children survived): all 18 had been transferred to an ICU for surveillance. Since patient care in PICUs is far more costly and manpower-intense than in general emergency rooms, we propose that a protocol of 6 h of surveillance in the emergency department is adequate and safe for most children who had been stung by LQS. Only children who develop systemic manifestations should be hospitalized and transferred to the intensive care unit. (5) Further prospective studies should be conducted to define specific subgroups that may benefit from these recommendations.

Adolescent↗

Regulation of UCP3 by nucleotides is different from regulation of UCP1.

UCP3 is an isoform of UCP1, expressed primarily in skeletal muscle. Functional properties of UCP3 are still largely unknown. Here, we report about the expression of UCP3 and of UCP1 in inclusion bodies of Escherichia coli. On solubilization and reconstitution into proteoliposomes, both UCP3 and UCP1 transport Cl- at rates equal to the reconstituted native UCP1. Cl- transport is inhibited by low concentrations of ATP, ADP, GTP and GDP. However, no H+ transport activity is found possibly due to the lack of a cofactor presents in UCP from mitochondria. The specificity of inhibition by nucleoside tri- and diphosphate is different between UCP1 and UCP3. UCP1 is more sensitive to tri- than diphosphate whereas in UCP3, the gradient is reverse. These results show a new paradigm for the regulation of thermogenesis at various tissues by the ATP/ADP ratio. In brown adipose tissue, the thermogenesis is correlated with a low ATP/ADP whereas in skeletal muscle, non-shivering thermogenesis is active at a high ATP/ADP ratio, i.e. in the resting state.

Adipose Tissue↗