ICln is a ubiquitously expressed eukaryotic protein. Expression of the protein in Xenopus laevis oocytes, the knocking-down of the protein in fibroblasts, or the reconstitution of the protein in lipid bilayer led to the assumption that... more
ICln is a ubiquitously expressed eukaryotic protein. Expression of the protein in Xenopus laevis oocytes, the knocking-down of the protein in fibroblasts, or the reconstitution of the protein in lipid bilayer led to the assumption that this protein is an ionic channel or a significant part thereof. However, other possible roles for ICln in potential regulatory mechanisms have been postulated, as diverse as regulator of cell morphology by interacting with the Skb1 protein and/or interaction with core spliceosomal proteins. Here we show that ICln is able to interact with SnRNP core proteins SmD1, SmD2, SmD3, SmX5 and SmB/B'.
Research Interests: Chemistry, Cell Biology, Ion Channels, Saccharomyces cerevisiae, Protein-Protein Interaction, and 14 moreXenopus, Cell morphology, Humans, Animals, Proteins, Medical Physiology, Xenopus laevis, Protein Expression, Yeast Two-hybrid System, Amino Acid Sequence, Base Sequence, Protein Binding, Lipid Bilayer, and Molecular Sequence Data
Distal forearm fractures in younger adults are rare injuries resulting from high energy trauma. Treatment options vary from cast fixation, external fixator, percutaneus pinning and open reduction and internal fixation. We retrospectively... more
Distal forearm fractures in younger adults are rare injuries resulting from high energy trauma. Treatment options vary from cast fixation, external fixator, percutaneus pinning and open reduction and internal fixation. We retrospectively reviewed 13 patients aged 18-59 from 1996 to 2005 with a distal unstable forearm fracture. All were treated with open reduction and internal fixation of the radius. The ulna was stabilized either by an open reduction and internal fixation or by a closed reduction with or without pin fixation and cast fixation in all cases. At follow-up, we evaluated the radiologic results in terms of forearm fracture retention and functional outcome according to the wrist score by Krimmer. Radial inclination amounted to 24 degrees at the injured side when compared to 27 degrees at the non-injured side, palmar tilt was 3 degrees versus 7 degrees and ulna variance was -2 versus -1 mm. According to the modified wrist score by Krimmer, seven excellent, two good and four fair results were achieved. The range of motion of the injured wrist joint was 149 degrees of rotation, in the sagittal plane 106 degrees , frontal plane 61 degrees and on the non-injured side rotation was 171 degrees , and movement in the sagittal plane was 146 degrees and 79 degrees in the frontal plane. Decreased forearm rotation (107 degrees vs. 162 degrees ) and decreased range of motion in the sagittal plane (77 degrees vs. 114 degrees ) were measured in patient following open reduction and internal fixation of radius and ulna compared to the outcome in patients with open reduction and internal fixation of the radius and closed reduction of the ulna. Grip strength of the injured side averaged 350 N versus 440 N which is 76% of that of the opposite side. All patients stated no pain at rest and some experienced slight pain at work. Three patients had an excellent performance at daily activities, nine patients presented problems with certain activities, and one patient showed severe limitations. Open reduction and internal fixation of the radius is the keystone in treating distal forearm fracture. In case of stable retention of the ulnar head after closed reduction, cast fixation with or without percutaneus pin fixation is a sufficient method to treat unstable distal forearm fractures. In patients with remaining instability of the distal ulna fracture, ORIF is indicated.
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Proximal humerus fractures remain challenging especially in the elderly. Biomechanical data put semi-rigid implants in favour of osteopenic or osteoporotic situation. Little surgical side damage is associated with a minimal invasive... more
Proximal humerus fractures remain challenging especially in the elderly. Biomechanical data put semi-rigid implants in favour of osteopenic or osteoporotic situation. Little surgical side damage is associated with a minimal invasive approach of these implants. The aim of this study was to evaluate the mechanical properties of three such implants. Fresh frozen cadaver specimens were mounted as proposed by the distributors. Three different implants were used: LCP-PH (locking compression plate proximal humerus, Synthes, Austria), HB (humerus block, Synthes, Austria), and IMC (intramedullary claw, ITS, Austria). Subcapital fracture was simulated by resecting a 5 mm gap. All specimens were comparable in "B" (one), "M" (ineral) and "D" (ensity). Four load cases were tested: varus bending, medial shearing and axial torque. A cyclic test (1,000 cycles) was performed in the first load case (varus stress) for all three implants. The LCP-PH was the most rigid in all three load cases, always followed by the HB. The IMC was the most elastic device with almost immeasurable values in axial torque. In the cyclic setting, the load reduction of the HB followed by the LCP-PH was significantly better than that for the IMC. The differences in stiffness are varying tremendously. The IMC is the implant with the lowest stiffness in all load cases and the highest load reduction. New "semi-rigids" claim good clinical performance, yet prospective clinical studies have to prove this. It is unlikely that the IMC can maintain fracture reduction in fracture situations of complex nature (no ligamentotaxis).
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Research Interests:
NIH 3T3 fibroblasts expressing the ras oncogene (+ras cells) respond to bradykinin, bombesin or serum with sustained oscillations of cell membrane potential reflecting oscillations of intracellular calcium activity and subsequent... more
NIH 3T3 fibroblasts expressing the ras oncogene (+ras cells) respond to bradykinin, bombesin or serum with sustained oscillations of cell membrane potential reflecting oscillations of intracellular calcium activity and subsequent activation of calcium-sensitive K+ channels. In contrast, identical cells not expressing the oncogene (-ras cells) respond to bradykinin with a single, transient hyperpolarization of the cell membrane. Furthermore, +ras cells are characterized by a serum-independent proliferation, an increase in cell volume and a marked reorganization of the cytoskeleton. It has been shown previously that the calcium channel blocker nifedipine, but not verapamil and diltiazem, inhibits oscillations of cell membrane potential as well as proliferation. In this study, we have examined the effect of several calcium channel blockers (bepridil, nifedipine, verapamil, diltiazem) on the proliferation, volume and cytoskeletal reorganization of +ras cells. Bepridil (10 mumol/l), whic...
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It is not resolved whether the anionic channel involved in volume regulation after cell swelling comprises one or more subunits. Moreover, it remains to be determined which of the different proteins cloned so far, for which an involvement... more
It is not resolved whether the anionic channel involved in volume regulation after cell swelling comprises one or more subunits. Moreover, it remains to be determined which of the different proteins cloned so far, for which an involvement in cell volume regulation has been postulated, is the ideal candidate. In this review, we consider the role of the ICln protein, cloned from MDCK cells, in cell volume regulation.
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Research Interests: Molecular Biology, Biology, Biological Chemistry, Ion Channels, Biological Sciences, and 15 moreCell line, Humans, Sequence alignment, Animals, Biological, Promoter, Proteins, CHEMICAL SCIENCES, Transfection, Gene Expression Regulation, Cell Size, Molecular Sequence Data, binding sites, Haplorhini, and Medical and Health Sciences
ICln is a ubiquitously expressed eukaryotic protein. Expression of the protein in Xenopus laevis oocytes, the knocking-down of the protein in fibroblasts, or the reconstitution of the protein in lipid bilayer led to the assumption that... more
ICln is a ubiquitously expressed eukaryotic protein. Expression of the protein in Xenopus laevis oocytes, the knocking-down of the protein in fibroblasts, or the reconstitution of the protein in lipid bilayer led to the assumption that this protein is an ionic channel or a significant part thereof. However, other possible roles for ICln in potential regulatory mechanisms have been postulated, as diverse as regulator of cell morphology by interacting with the Skb1 protein and/or interaction with core spliceosomal proteins. Here we show that ICln is able to interact with SnRNP core proteins SmD1, SmD2, SmD3, SmX5 and SmB/B'.
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Research Interests: Adolescent, Medicine, Probability, Humans, Child, and 15 moreFemale, Male, Dislocations, Fracture Healing, Clinical Sciences, Retrospective Studies, Age Factors, Bone remodeling, Risk Assessment, Retrospective Study, Growth Plate, Injury Severity Score, Child preschool, Pronation, and functional outcome
Distal forearm fractures in younger adults are rare injuries resulting from high energy trauma. Treatment options vary from cast fixation, external fixator, percutaneus pinning and open reduction and internal fixation. We retrospectively... more
Distal forearm fractures in younger adults are rare injuries resulting from high energy trauma. Treatment options vary from cast fixation, external fixator, percutaneus pinning and open reduction and internal fixation. We retrospectively reviewed 13 patients aged 18-59 from 1996 to 2005 with a distal unstable forearm fracture. All were treated with open reduction and internal fixation of the radius. The ulna was stabilized either by an open reduction and internal fixation or by a closed reduction with or without pin fixation and cast fixation in all cases. At follow-up, we evaluated the radiologic results in terms of forearm fracture retention and functional outcome according to the wrist score by Krimmer. Radial inclination amounted to 24 degrees at the injured side when compared to 27 degrees at the non-injured side, palmar tilt was 3 degrees versus 7 degrees and ulna variance was -2 versus -1 mm. According to the modified wrist score by Krimmer, seven excellent, two good and four fair results were achieved. The range of motion of the injured wrist joint was 149 degrees of rotation, in the sagittal plane 106 degrees , frontal plane 61 degrees and on the non-injured side rotation was 171 degrees , and movement in the sagittal plane was 146 degrees and 79 degrees in the frontal plane. Decreased forearm rotation (107 degrees vs. 162 degrees ) and decreased range of motion in the sagittal plane (77 degrees vs. 114 degrees ) were measured in patient following open reduction and internal fixation of radius and ulna compared to the outcome in patients with open reduction and internal fixation of the radius and closed reduction of the ulna. Grip strength of the injured side averaged 350 N versus 440 N which is 76% of that of the opposite side. All patients stated no pain at rest and some experienced slight pain at work. Three patients had an excellent performance at daily activities, nine patients presented problems with certain activities, and one patient showed severe limitations. Open reduction and internal fixation of the radius is the keystone in treating distal forearm fracture. In case of stable retention of the ulnar head after closed reduction, cast fixation with or without percutaneus pin fixation is a sufficient method to treat unstable distal forearm fractures. In patients with remaining instability of the distal ulna fracture, ORIF is indicated.
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Research Interests: Orthopedic Surgery, Medicine, Prospective studies, Humans, Female, and 15 moreMale, Cost effectiveness, Clinical Sciences, Middle Aged, Adult, Minimally Invasive, Immobilization, Prospective Study, Orthopedic Procedures, Conservative treatment, Grip strength, Bone screws, Costs and Cost Analysis, Indirect Cost, and Fractures Bone
Proximal humerus fractures remain challenging especially in the elderly. Biomechanical data put semi-rigid implants in favour of osteopenic or osteoporotic situation. Little surgical side damage is associated with a minimal invasive... more
Proximal humerus fractures remain challenging especially in the elderly. Biomechanical data put semi-rigid implants in favour of osteopenic or osteoporotic situation. Little surgical side damage is associated with a minimal invasive approach of these implants. The aim of this study was to evaluate the mechanical properties of three such implants. Fresh frozen cadaver specimens were mounted as proposed by the distributors. Three different implants were used: LCP-PH (locking compression plate proximal humerus, Synthes, Austria), HB (humerus block, Synthes, Austria), and IMC (intramedullary claw, ITS, Austria). Subcapital fracture was simulated by resecting a 5 mm gap. All specimens were comparable in "B" (one), "M" (ineral) and "D" (ensity). Four load cases were tested: varus bending, medial shearing and axial torque. A cyclic test (1,000 cycles) was performed in the first load case (varus stress) for all three implants. The LCP-PH was the most rigid in all three load cases, always followed by the HB. The IMC was the most elastic device with almost immeasurable values in axial torque. In the cyclic setting, the load reduction of the HB followed by the LCP-PH was significantly better than that for the IMC. The differences in stiffness are varying tremendously. The IMC is the implant with the lowest stiffness in all load cases and the highest load reduction. New "semi-rigids" claim good clinical performance, yet prospective clinical studies have to prove this. It is unlikely that the IMC can maintain fracture reduction in fracture situations of complex nature (no ligamentotaxis).
Research Interests:
Research Interests:
Dear Editor, The setup for this selective testing of implants was chosen for two reasons. First, we wanted to compare our results with those of an existing study which had tested the LCPPH using the same setup [1]. Second, we believe that... more
Dear Editor, The setup for this selective testing of implants was chosen for two reasons. First, we wanted to compare our results with those of an existing study which had tested the LCPPH using the same setup [1]. Second, we believe that this selective setup is the most appropriate one to reveal the biomechanical performance of implants on the proximal humerus. As stated, we do think that there are fracture situations with interfragmentary ligamentotaxis that are relatively stable after reduction. This is the type of fracture that was treated traditionally with percutaneous pins or more recently with the Humerus Block with good results. These fracture situations are not the subject of our study. In our testing, the IMC showed the lowest stiVness in all cases of load. Subsequently, throughout cyclic testing, it failed to maintain even the initially modest stiVness and showed the greatest loss in load reduction. Therefore, we concluded that it appears highly questionable that the IMC should be capable of stabilizing complex, highly unstable situations without interfragmentary ligamentotaxis. In an ethic commission approved prospective randomised clinical trial testing IMC versus HB, we treated severely dislocated isolated two-part (subcapital) proximal humerus fractures. Inclusion criteria were dislocation of more than half of the width of the shaft or an angulation of more than 45° between the head and the shaft fragment. Patients were randomly assigned to one of the two implants. In both groups all patients were female; mean age was comparable in both groups (70 years IMC/ 68 years HB). In this series of largely geriatric fractures (with concomitant poor bone quality), the IMC was not capable of Wxing the intraoperatively impacted fracture site in an acceptable position until healing had occurred. We stopped the study because of the high rate of severe complications of the IMC implant. About 60% of the IMC treated patients suVered severe, implant-related complications. To sum up, the IMC implant may work in relatively stable subcapital fractures with intact ligamentotaxis and little displacement. These fractures, according to our protocol, are mostly treated nonoperatively. However, in unstable situations with the aforementioned criteria, we failed to demonstrate suYcient holding strength of the implant as well in the biomechnical setup as in the clinical situation.
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Cell volume regulation & a ubiquitous cell regulatory mechanism based on meticulously controlled ion transport mechanisms. Keeping the absolute volume constant seems to be of the highest priority for most cells and is achieved at the... more
Cell volume regulation & a ubiquitous cell regulatory mechanism based on meticulously controlled ion transport mechanisms. Keeping the absolute volume constant seems to be of the highest priority for most cells and is achieved at the expense of altered intracellular ion concentrations. We have been able to demonstrate that Ic# ~ a chloride channel cloned from epithelial cells, is paramount for the ability of" swollen cells to regulate their volume back to that under resting conditions. A unique feature of Icl n is the distinct sensitivity of these channels' for nucleotides and nucleoside analogues added to the extracellular fluid. In addition, cromolyn sodium and nedocromil sodium, drug, s used by patients with asthma, are able to impede the .function of these channels. (J Allergy Clin Immunol 1996;98:$98-10L)
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To evaluate the sequelae of distal intraarticular radius fracture with regard to the development of arthritis and clinical symptoms. In a retrospective follow-up examination, 72 patients with a distal intraarticular radius fracture could... more
To evaluate the sequelae of distal intraarticular radius fracture with regard to the development of arthritis and clinical symptoms. In a retrospective follow-up examination, 72 patients with a distal intraarticular radius fracture could be included for clinical and radiological investigation 9 years following the trauma. All fractures were treated by ORIF and cortico-cancellous bone grafting. Radiological evaluation revealed 5.1 degrees palmar tilt, 19.1 degrees radial tilt and the ulnar variance amounted to -0.5 mm. The articular cavity depth in the sagittal plane measured with 4.6 mm, 1.2 mm more than the non-involved side. Articular step-off was noticed in 6 patients. According to the Knirk and Jupiter classification system, two patients healed without arthritis, 43 patients presented arthritis stage 1, and 27 stage 2. Evaluation of the data showed a significant correlation between arthritis and articular cavity depth. But arthritis had neither influence on the DASH, nor the pain level. On the other hand, arthritis led to decreased sagittal wrist motion. ORIF of distal intraarticular radius fractures led to predictable results concerning restoration of length and form of the distal radius. Arthritis had a minor influence on the clinical end result.
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The study was made to evaluate the role of MR imaging in pediatric distal forearm fractures by comparison with the findings of plain radiographs and MRI. 38 patients (27 boys and 11 girls, mean age of 12 years, range 7 to 15 years) with... more
The study was made to evaluate the role of MR imaging in pediatric distal forearm fractures by comparison with the findings of plain radiographs and MRI. 38 patients (27 boys and 11 girls, mean age of 12 years, range 7 to 15 years) with radiographically open distal radius and ulna growth plates requiring first aid for a fracture of the distal third of the forearm, were included in this study. The fractures were diagnosed on plain radiographs and conservative treatment was performed. In 35 patients MR imaging was performed within 3 weeks after the accident and in 3 patients MRI was performed after 6 to 9 weeks because of persistent wrist pain. Fifteen Salter/Harris II injuries of the radius and 1 of the ulna, 1 torus fracture of the radius and 2 of the ulna, 12 greenstick fractures of the radius and 3 of the ulna, 10 complete displaced radius fractures and 15 ulnar styloid fractures were found on plain radiographs. Twelve patients had evidence of associated triangular fibrocartilage ...
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To evaluate the sequelae of distal intraarticular radius fracture with regard to the development of arthritis and clinical symptoms. In a retrospective follow-up examination, 72 patients with a distal intraarticular radius fracture could... more
To evaluate the sequelae of distal intraarticular radius fracture with regard to the development of arthritis and clinical symptoms. In a retrospective follow-up examination, 72 patients with a distal intraarticular radius fracture could be included for clinical and radiological investigation 9 years following the trauma. All fractures were treated by ORIF and cortico-cancellous bone grafting. Radiological evaluation revealed 5.1 degrees palmar tilt, 19.1 degrees radial tilt and the ulnar variance amounted to -0.5 mm. The articular cavity depth in the sagittal plane measured with 4.6 mm, 1.2 mm more than the non-involved side. Articular step-off was noticed in 6 patients. According to the Knirk and Jupiter classification system, two patients healed without arthritis, 43 patients presented arthritis stage 1, and 27 stage 2. Evaluation of the data showed a significant correlation between arthritis and articular cavity depth. But arthritis had neither influence on the DASH, nor the pai...
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The present study was designed to evaluate long-term outcome of upper extremities and subjective self-assessment of patient disability after a Kapandji-Sauvé procedure by means of the DASH score. Between 1986 and 1996, a modified... more
The present study was designed to evaluate long-term outcome of upper extremities and subjective self-assessment of patient disability after a Kapandji-Sauvé procedure by means of the DASH score. Between 1986 and 1996, a modified Kapandji-Sauvé procedure was performed in 117 patients with painfully limited forearm rotation and arthrosis of the distal radioulnar joint (DRUJ). Of the 117 patients, 73 women and 32 men, whose ages at operation ranged from 22 to 74 years (average, 58 years) were retrospectively reviewed clinically and radiologically eight years (range, five to twelve years) after the operation. The DASH questionnaire was used in 43 patients. The mean DASH score was 28 points (range, 0 to 53 points). The mean score in part A was 1.9 points, in part B 1.8 points. Worst outcomes were noted for activities requiring the exertion of force. Pain was reduced in 97 % of the patients. Forearm rotation and grip strength improved in all patients. Our clinical findings suggest that t...
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Distal radius fracture often presents a metaphyseal void which is more extended in elderly, osteoporotic patients. Bone graft and bone substitutes are reported to be beneficial in maintaining metaphyseal reduction. We performed a... more
Distal radius fracture often presents a metaphyseal void which is more extended in elderly, osteoporotic patients. Bone graft and bone substitutes are reported to be beneficial in maintaining metaphyseal reduction. We performed a prospective study on 52 menopausal, osteoporotic women with unstable intra-articular distal radius fractures to compare the outcome of percutaneous pinning and immobilisation in a cast for 6 weeks with that using injectable calcium phosphate bone cement (Norian Skeletal Repair System, SRS) to supplement pin and screw fixation and immobilisation in a cast for 3 weeks. All patients were reviewed 2 years (range 21-29 months) after surgery. Patients treated with SRS had better functional outcome, restoration of movement and grip strength ( p<0.001). In this group there was 1 mm loss of radial length, 3 degrees loss of radial inclination and 7 degrees loss of palmar tilt. In the control group radial length decreased 3 mm, radial inclination decreased 11 degre...
Research Interests: Orthopedic Surgery, Fracture, Probability, Bone graft, Prospective studies, and 15 moreHumans, Female, Calcium Phosphates, Bone Cement, Fracture Healing, Clinical Sciences, Biomaterial, Association, Aged, Middle Aged, Fixation, Calcium Phosphate, Bone Cements, Grip strength, and Injury Severity Score
Osteoporotic fractures of the proximal humerus show an increasing incidence. Osteoporosis not only influences the fracture risk after low-energy trauma, but also affects the mechanical stability of internal fixation. Preoperative... more
Osteoporotic fractures of the proximal humerus show an increasing incidence. Osteoporosis not only influences the fracture risk after low-energy trauma, but also affects the mechanical stability of internal fixation. Preoperative assessment of the local bone quality may be useful in the surgical treatment of patients sustaining these injuries. The aim of the present study was to present a method for the preoperative assessment of the local cancellous bone mineral density (BMD) of the proximal humerus using CT data. In the first part of the study, CT scans of 30 patients with unilateral fractures of the proximal humerus after low-energy trauma were used. The local BMD was assessed on the contralateral uninjured side. All 30 patients additionally underwent dual-emission X-ray absorptiometry (DXA) of the lumbar spine, proximal femur, and forearm of the side of the uninjured proximal humerus within 6 weeks after trauma. Three independent trauma surgeons performed measurements on the uninjured proximal humerus twice with a time interval of 4 weeks in order to assess the inter- and intraobserver reliability of the method. In the second part of the study, the local BMD of 507 patients with either proximal humerus fractures or chronic shoulder instability was assessed by a single trauma surgeon. In both parts, the average HU values in standardized ROIs of the humeral head were automatically calculated after correcting for HU values below the water equivalent. A linear calibration equation was computed for the calculation from HU to BMD using a calibration device (EFP). The intra- and interobserver reliability was high (ICC &amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;gt; 0.95). Correlation coefficients between the local BMD of the proximal humerus and other anatomical sites were between 0.35 (lumbar spine) and 0.64 (forearm). We found a high correlation between the local BMD and age. The BMD in the fracture group was significantly lower than in the instability group. These patients were significantly older and more likely to be female. Our method may provide a preoperative tool for the assessment of the local bone quality of the proximal humerus using CT data. Therapeutic adjustments such as augmentation or primary arthroplasty may be considered in patients with very low local BMD.
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Research Interests:
Fluorescence-optical measurements of the intracellular chloride concentration facilitate identification of chloride movements across the cell membrane of living cells. The two main dyes used for this purpose are... more
Fluorescence-optical measurements of the intracellular chloride concentration facilitate identification of chloride movements across the cell membrane of living cells. The two main dyes used for this purpose are 6-methoxy-N-(3-sulfopropyl)quinolinium (SPQ) and 6-methoxy-quinolyl acetoethyl ester (MQAE). The use of both substances is impaired by their poor membrane permeability and therefore limited loading of the cells to be studied. Here we report the use of 6-methoxy-N-ethylquinolinium iodide (MEQ), a chloride-sensitive dye for which a membrane-permeable form is easily prepared. This makes the loading procedure as easy as with the acetoxymethyl (AM) forms of other dyes for sensing intracellular ions. In addition, the original method, which described absolute concentration measurements of chloride in the cytosol, was modified in so far as only relative measurements were made. This avoids the known limitations of single wavelength excitation and emission dyes with respect to exact concentration measurements. Moreover, to enhance the signal-to-noise ratio the driving force for chloride was considerably increased by changing the original direction of the anion flux in the cells under investigation. We verified the method by using fibroblasts and activating ICln, a putative chloride channel cloned from epithelial cells and of paramount importance in the regulatory volume decrease in these cells. In the presence of SCN- the MEQ quench measured in NIH 3T3 fibroblasts is dramatically enhanced in hypotonically challenged cells compared with cells under isotonic conditions. Antisense oligodeoxynucleotides sensing ICln considerably impeded the swelling-induced chloride current (ICl) in NIH 3T3 fibroblasts. Accordingly, the chloride movement measured by the SCN- quench of the MEQ signal was significantly reduced. Similar results can be obtained in the presence of 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) or 4, 4&#39;-diisothiocyanatostilbene-2,2&#39;-disulfonic acid (DIDS), two known blockers of chloride transport in the plasma membrane of a variety of cells. In conclusion, fluroscence-optical measurements using MEQ as the chloride-sensitive dye provide a reliable and easy-to-use method for measuring changes of the chloride flux across the cell membrane of living cells.
Research Interests: Physiology, Electrophysiology, Patch-clamp and imaging techniques, Mice, Animals, and 14 moreT cells, Medical Physiology, European, Benzoic Acid, Chloride Channel, Signal to Noise Ratio, Epithelial Cell Biology, Plasma Membrane, Regulatory volume decrease, Cell Size, Chlorides, Driving force, Cell Membrane, and fluorescent dyes
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Cell volume regulation is an essential feature of most cells. After swelling in hypotonic media, the simultaneous activation of potassium and chloride channels is believed to be the initial, time-determining step in cell volume... more
Cell volume regulation is an essential feature of most cells. After swelling in hypotonic media, the simultaneous activation of potassium and chloride channels is believed to be the initial, time-determining step in cell volume regulation. The activation of both pathways is functionally linked and enables the cells to lose ions and water, subsequently leading to cell shrinkage and readjustment of the initial volume. NIH 3T3 fibroblasts efficiently regulate their volume after swelling and bear chloride channels that are activated by decreasing extracellular osmolarity. The chloride current elicited in these cells after swelling is reminiscent of the current found in oocytes expressing an outwardly rectifying chloride current termed ICln. Introduction of antisense oligodeoxynucleotides complementary to the first 30 nucleotides of the coding region of the ICln channel into NIH 3T3 fibroblasts suppresses the activation of the swelling-induced chloride current. The experiments directly demonstrate an unambiguous link between a volume-activated chloride current and a cloned protein involved in chloride transport.
Research Interests: Physiology, Kinetics, Patch-clamp and imaging techniques, Mice, Animals, and 15 morePotassium, Medical Physiology, European, Epithelium, Epithelial cells, Chloride Channel, Antisense Oligonucleotide Drugs, Biotransformation, Base Sequence, Regulatory volume decrease, Cell Volume, Nucleotides, Cell Size, Molecular Sequence Data, and fibroblasts
Reconstitution of purified ICln in lipid bilayer leads to functional ion channels showing varying rectification. The reconstituted single channels have a conductance of approximately equal to 3 pS and their open probability is sensitive... more
Reconstitution of purified ICln in lipid bilayer leads to functional ion channels showing varying rectification. The reconstituted single channels have a conductance of approximately equal to 3 pS and their open probability is sensitive to nucleoside analogues. Mutation of a putative nucleotide binding site identified at the predicted extracellular mouth of the ICln channel protein leads to the reduction of the nucleoside-analogue sensitivity. Reconstituted ICln channels can be permeated both by cations and anions. The relative permeability of cations over anions depends on the presence of calcium. In the presence of calcium reconstituted ICln channels are more permeable to bromide than chloride, and more permeable to potassium than sodium. Similarly in NIH3T3 fibroblasts, the relative permeability of cations over anions of swelling-dependent chloride channels depends on extracellular calcium. Site-directed mutagenesis revealed the calcium-binding site responsible for the shift of the selectivity from cations towards anions of reconstituted ICln channels. Additional indirect structural information has been obtained by mutating a histidine in the predicted pore region of ICln. This histidine seems to have access to the ion-conducting tunnel of the pore. Our experiments show that ICln can act as an ionic channel, which does not exclude additional functions of the protein in regulatory mechanisms of the cell. Since knocking down the ICln protein in fibroblasts and epithelial cells leads to an impaired regulatory volume decrease (RVD) after cytoplasmic swelling and reconstituted ICln channels show several biophysical features of ion channels activated after swelling, ICln is a molecular candidate for these channels.
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Research Interests: Protein Folding, Biological Chemistry, Ion Channels, Patch-clamp and imaging techniques, Biological Sciences, and 12 moreDogs, Humans, Protein Kinases, Mice, Animals, Biological, Phosphorylation, CHEMICAL SCIENCES, Amino Acid Sequence, Blood Proteins, Molecular Sequence Data, and Medical and Health Sciences
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Research Interests: Biophysics, Kinetics, Immunohistochemistry, Biological Chemistry, Ion Channels, and 15 moreFluorescence Resonance Energy Transfer, Biological Sciences, Dogs, Cell line, Kidney, Animals, Male, Biological, CHEMICAL SCIENCES, Anions, Lipid bilayers, Human Fibroblasts, Ions, Cell Membrane, and fibroblasts
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Expression cloning revealed a chloride channel (ICln) that we found to be fundamental for the regulatory volume decrease in a variety of cells. The chromosomal localization of the human ICln-gene showed two loci, one at chromosome 11 in... more
Expression cloning revealed a chloride channel (ICln) that we found to be fundamental for the regulatory volume decrease in a variety of cells. The chromosomal localization of the human ICln-gene showed two loci, one at chromosome 11 in position q13.5-q14.1, termed CLNS1A, and a second one at chromosome 6 at position p12.1-q13, termed CLNS1B. In this study, we offer a detailed characterization of the CLNS1A gene and provide the exact position (6p12) and sequence data of CLNS1B, an intronless gene 91.3% homologous to the coding region of CLNS1A.
Research Interests: Genetics, Ion Channels, Humans, Fluorescence in situ hybridization, Sequence alignment, and 14 morePolymerase Chain Reaction, Gene, Introns, Fluorescent in situ hybridization, Benzoic Acid, Chloride Channel, Genomic Library, Amino Acid Sequence, Base Sequence, Open Reading Frame, Reverse Transcriptase, Regulatory volume decrease, Nucleotides, and Molecular Sequence Data
In previous studies we have shown that the expression of the transforming Ha-ras oncogene in NIH 3T3 fibroblasts stimulates cellular calcium entry, which triggers oscillatory calcium induced calcium release from internal stores. The... more
In previous studies we have shown that the expression of the transforming Ha-ras oncogene in NIH 3T3 fibroblasts stimulates cellular calcium entry, which triggers oscillatory calcium induced calcium release from internal stores. The intracellular calcium oscillations lead to cytoskeletal remodeling by actin stress fiber depolymerization and activation of the Na(+)/H(+) exchanger thus mediating cell swelling and intracellular alkalosis, both important mitogenic signals. This is evidenced by abrogation of Ha-ras induced growth factor independent cell proliferation by interference with any of these events, i.e. by inhibition of cellular calcium entry or inhibition of the Na(+)/H(+) exchanger. As shown in this study, simvastatin, an inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase, the key enzyme for cholesterol biosynthesis, is able to prevent these events following the expression of the transforming Ha-ras oncogene. We show, that simvastatin inhibits farnesylation dependent membrane translocation of a CAAX motive bearing yellow fluorescent protein and suppresses Ha-ras stimulated cellular calcium influx, which can be identified as capacitative calcium entry. In addition simvastatin is able to block regulatory volume decrease channels and to suppress the cytoskeletal remodeling, intracellular alkalinization, increase in cell volume and growth factor independent cell proliferation induced by the oncogene. Thus simvastatin is able to prevent crucial cellular events following expression of the transforming Ha-ras oncogene.