Biomedical subjects

A Moroni

Publications and source records attributed to A Moroni.

At least 37 records · Page 2Linked to original sources

PubMed2000

Kinetic and ionic properties of the human HCN2 pacemaker channel.

Human cDNA coding for the hyperpolarization-activated "pacemaker" channel HCN2 was expressed in Phoenix cells and yielded an inward current (IhHCN2) activated on hyperpolarization. The average IhHCN2 was half-activated at -83.1 mV and its kinetics could be described by second-order Hodgkin-Huxley gating. The time constant curve was bell-shaped and peaked at -82.2 mV. With 115 mM external Na+ and 30 mM external K+, IhHCN2 reversed at -17.1 mV, and had a mean conductance of 5.6 nS. Reducing the external K+ or Na+ concentration led to a concentration-dependent reduction of the IhHCN2 conductance and to a hyperpolarizing shift of reversal potential. External Cs+ ions (5 mM) blocked IhHCN2 in a voltage-dependent way according to a Woodhull-type block model, at an electrical distance of 0.66 from the external membrane surface, and with a dissociation constant of 15 mM at 0 mV. Increasing cytoplasmic cAMP using forskolin increased IhHCN2 by shifting the current activation curve to more positive voltages (11.7 mV). Exposure of the intracellular side of inside-out macro-patches to cAMP led to a depolarizing shift of the channel open probability curve (15.2 mV with 10 microM cAMP). These results indicate that although hHCN2 channels share several properties with native cardiac f-channels, differences also exist in permeability and block properties, suggesting that native channels may not be composed simply of homomeric constructs.

Biological Clocks↗

PubMed2000

Mutation in pore domain uncovers cation- and voltage-sensitive recovery from inactivation in KAT1 channel.

Effects of threonine substitution by glutamine at position 256 in the pore of the KAT1 channel have been investigated by voltage-clamp, using heterologous gene expression in Xenopus oocytes. The major discrepancy in T256Q from the wild-type channel (wt) was cation specific. While K(+) currents were reduced in a largely scalar fashion, the NH(4)(+) current exhibited slow, voltage-dependent inhibition during hyperpolarization. The same effects could be induced in wt, or intensified in T256Q, by addition of the impermeant cation methylammonium (MA(+)) to the bath. This stresses that both the mutation and MA(+) affect a mechanism already present in the wt. Assuming that current inhibition could be described as entry of the channel into an inactive state, we modeled in both wt and in T256Q the relaxation kinetics of the clamp currents by a C-O-I gating scheme, where C (closed) and I (inactivated) are nonconductive states, and O is an open state allowing K(+) and NH(4)(+) passage. The key reaction is the transition I-O. This cation-sensitive transition step ensures release of the channel from the inactive state and is approximately 30 times smaller in T256Q compared to wt. It can be inhibited by external MA(+) and is stimulated strongly by K(+) and weakly by NH(4)(+). This sensitivity of gating to external cations may prevent K(+) leakage from cation-starved cells.

Amino Acid Substitution↗

PubMed2000

Risk factors for intraoperative femoral fractures during total hip replacement.

BACKGROUND AND AIMS: Intraoperative femoral fractures are a serious complication of total hip replacement. The purpose of this study was to evaluate the risk factors of intraoperative femoral fractures in a retrospective analysis of a series of 3,566 total hip replacements. MATERIALS AND METHODS: The patients were divided into two groups, A and B. Group A patients had no intraoperative femoral fractures and Group B patients had intraoperative femoral fractures. In Group A there were 3,483 patients (97.7%) and in Group B, 83 (2.3%). The following potential risk factors were evaluated: sex, age, diagnosis, previous surgery at the homolateral hip, surgical approach, fixation type of the femoral component, prosthesis type, surgical stage during which the fracture occurred, and the lead operating surgeon. RESULTS: The fracture incidence was higher in females (p < 0.005) in uncemented femoral components (p = 0.005), in patients who had previous surgery at the homolateral hip (p < 0.005), and in revision surgery (p < 0.005). CONCLUSION: The analysis of intraoperative femoral fracture risk factors should allow the surgeon to improve the surgical performance and therefore reduce the incidence of this severe intraoperative complication.

Adult↗

PubMed1999

Characterization and properties of dominant-negative mutants of the ras-specific guanine nucleotide exchange factor CDC25(Mm).

Ras proteins are small GTPases playing a pivotal role in cell proliferation and differentiation. Their activation depends on the competing action of GTPase activating proteins and guanine nucleotide exchange factors (GEF). The properties of two dominant-negative mutants within the catalytic domains of the ras-specific GEF, CDC25(Mm), are described. In vitro, the mutant GEF(W1056E) and GEF(T1184E) proteins are catalytically inactive, are able to efficiently displace wild-type GEF from p21(ras), and strongly reduce affinity of the nucleotide-free ras x GEF complex for the incoming nucleotide, thus resulting in the formation of a stable ras.GEF binary complex. Consistent with their in vitro properties, the two mutant GEFs bring about a dramatic reduction in ras-dependent fos-luciferase activity in mouse fibroblasts. The stable ectopic expression of the GEF(W1056E) mutant in smooth muscle cells effectively reduced growth rate and DNA synthesis with no detectable morphological changes.

Animals↗

PubMed1999

Action of internal pronase on the f-channel kinetics in the rabbit SA node.

1. The hyperpolarization-activated If current was recorded in inside-out macropatches from sino-atrial (SA) node myocytes during exposure of their intracellular side to pronase, in an attempt to verify if cytoplasmic f-channel domains are involved in both voltage- and cAMP-dependent gating. 2. Superfusion with pronase caused a quick, dramatic acceleration of channel opening upon hyperpolarization and slowing, rapidly progressing into full blockade, of channel closing upon depolarization; these changes persisted after wash off of pronase and were irreversible, indicating proteolytic cleavage of channel regions which contribute to gating. 3. If recorded from patches normally responding to cAMP became totally insensitive to cAMP following pronase treatment, indicating partial or total removal of channel regions involved in the cAMP-dependent activation. 4. The fully activated I-V relationship was not modified by pronase, indicating that internal proteolysis did not affect the f-channel conductance. 5. The changes in If kinetics induced by pronase were due to a large depolarizing shift of the f-channel open probability curve (56.5 +/- 1.1 mV, n = 7). 6. These results are consistent with the hypothesis that cytoplasmic f-channel regions are implicated in dual voltage- and cAMP-dependent gating; also, since pronase does not abolish hyperpolarization-activated opening, an intrinsic voltage-dependent gating mechanism must exist which is inaccessible to proteolytic cleavage. A model scheme able to account for these data thus includes an intrinsic gating mechanism operating at depolarized voltages, and a blocking mechanism coupled to cAMP binding to the channel.

Animals↗

PubMed1999

The human gene coding for HCN2, a pacemaker channel of the heart.

Hyperpolarization-activated, cyclic nucleotide-gated (HCN) channels, underlying 'pacemaker' currents (I(f)/Ih), are involved in pacemaker activity of cardiac sinoatrial node myocytes and central neurons. Several cDNAs deriving from four different genes were recently identified which code for channels characterized by six transmembrane domains and a cyclic nucleotide binding domain. We report here the identification of the human HCN2 gene and show that its functional expression in a human kidney cell line generates a current with properties similar to the native pacemaker f-channel of the heart. The hHCN2 gene maps to the telomeric region of chromosome 19, band p13.3. This is the first identification of a genetic locus coding for an HCN channel.

Amino Acid Sequence↗

PubMed1999

Measurement of insertion torque of tapered external fixation pins: A comparison between two experimental models.

The aim of this study was to compare an in vitro versus an ex vivo experimental model to test the insertion torque of two different types of external fixation pins. A torque measuring machine was developed in order to perform accurate measurements. Forty tapered pins made of stainless steel were utilized. Half of the pins were plasma-spray coated with hydroxyapatite (HA) and the other half remained uncoated. For the in vitro model 20 cylinders were used that were made of synthetic polymer according to ASTM standards. For the ex vivo model 10 fresh femora harvested from adult sheep were used. All the pins were implanted after predrilling, and insertion torque was measured. Statistical analysis of the in vitro versus the ex vivo model showed significant differences in both coated (p < 0. 0005) and uncoated (p = 0.002) external fixation pins. These results may be due to the surface roughness that caused significant friction between the HA coating and the polyvinylchloride in the in vitro model. The significant difference between the in vitro and ex vivo results lead us to state that the in vitro model does not realistically simulate the behavior of external fixation pins implanted in bone.

Animals↗

PubMed1999

Hydroxyapatite-coated orthopaedic screws as infection resistant materials: in vitro study.

The authors evaluated in vitro the adherence of a Staphylococcus epidermidis strain to hydroxyapatite-coated stainless-steel screws-used in orthopaedic surgery for external fracture fixation-in comparison with the adherence to uncoated screws. Evaluations were also performed on analogous groups of screws immersed for 72 and 168 h in a solution at 37 degrees C, in order to simulate the interstitial fluid in a simplified way. Adherence values on coated prostheses resulted significantly lower compared with those observed on metal prostheses, both in basal conditions and after immersion in saline solution. Moreover, both on coated and on uncoated screws a significant reduction in bacterial adherence was noted related to the duration of the prosthesis permanence in saline solution.

Bacterial Adhesion↗

PubMed1999

Hydroxyapatite fully coated conic hip prosthetic stem: a long term animal study.

The purpose of this paper is to evaluate in an animal model the long term results obtained with a prosthetic stem fully coated with hydroxyapatite. The cup was manufactured in polyethylene and was cemented. Six arthroplasties were performed in six sheep. After twelve months, the animals were euthanized and the femurs were harvested and processed for undecalcified sectioning. Twelve cross sections were cut perpendicularly to the longitudinal stem axis. Sections one to five corresponded to the area of the stem which, at the time of surgery, had a full initial contact between the bone and the prosthesis; sections six to ten corresponded to the area of the stem which, at the time of surgery, had a gap from 0 to 2 mm between the bone and the prosthesis; sections eleven and twelve had an initial gap larger than 2 mm. At one year after implantation, in the sections one to five, morphological analyses showed extensive direct contact between the bone and the hydroxyapatite coating. Bone prosthesis contact was lower in the sections six to ten. No contact was seen in sections eleven and twelve. Comparing bone to prosthesis contact of each subsequent section, from proximal to distal, the difference becomes significant with section five compared to section six (p < 0.00005). No detachment of the hydroxyapatite coating from the metallic substrate was observed in any section. In conclusion, this study shows that a conic shaped femoral stem, fully coated with hydroxyapatite gives very good histological and histomorphometric results at one year. Prosthesis osteointegration showed to be influenced by the initial bone to prosthesis contact. No direct bone to prosthesis contact was achieved if the initial bone to prosthesis gap was larger than 2 mm.

Animals↗

PubMed1998

Mutations at position 1122 in the catalytic domain of the mouse ras-specific guanine nucleotide exchange factor CDC25Mm originate both loss-of-function and gain-of-function proteins.

The role of two residues within the catalytic domain of CDC25Mm, a mouse ras-specific guanine nucleotide exchange factor (GEF), was investigated by site-directed mutagenesis. The function of the mutant proteins was tested in vivo in both a Saccharomyces cerevisiae cdc25 complementation assay and in a mammalian fos-luciferase assay, and in in vitro assays on human and yeast Ras proteins. Mutants CDC25Mm(E1048K) and CDC25Mm(S1122V) were shown to be (partly) inactive proteins, similar to their yeast homologs. Mutant CDC25Mm(S1122A) showed higher nucleotide exchange activity than the wild type protein on the basis of both in vitro and in vivo assays. Thus, alanine and valine substitutions at position 1122 within the GEF catalytic domain originate mutations with opposite biological properties, indicating an important role for position 1122 in GEF function.

3T3 Cells↗

PubMed1998

The impermeant ion methylammonium blocks K+ and NH4+ currents through KAT1 channel differently: evidence for ion interaction in channel permeation.

The permeation properties of KAT1, an inward rectifying potassium channel from plant cells, were investigated with different ions in the external medium. With either K+, NH4+ or methylammonium (MA) in the external solution, the channel, expressed in Xenopus oocytes, appeared permeable to K+ and, to a lesser extent, to NH4+ but not to the slightly bigger, methylated analogue of NH4+, MA. Substituting NH4+ for K+ shifted the voltage dependency of channel activation further negative and hastened activation kinetics. This suggests that channel operation depends on the transported substrate. In mixed solution (50 mM K+, 50 mM MA) MA inhibited K+ current in a voltage-independent manner. The maximum block did not exceed 50% of the K+ current. In contrast, when NH4+ was the permeant ion (50 mM NH4+, 50 mM MA) MA caused a voltage-dependent, slowly developing open channel block, achieving complete inhibition at very negative voltages. The latter block could be partially overcome by the addition of K+ in the external solution. The data support a model in which ions, after entering the channel pore, compete with different affinities for binding sites on their permeation pathway.

Ammonium Chloride↗

PubMed1998

The interface of bone microstructure and an innovative coating: an X-ray diffraction study.

The in vivo compatibility and degradation aspects of an innovative coating to be sprayed onto titanium implants were investigated. The surface of fluorinated apatite (fHA), consisting of fluorhydroxyapatite plasma sprayed in a vacuum atmosphere, was treated with carbonate to improve its biological compatibility. fHA coating was compared with titanium implants coated (a) with hydroxyapatite (HA) by the traditional plasma spraying, and (b) with titanium oxide (TiOx). Screw-shaped implants were inserted in the cortical bone of sheep tibiae. X-ray diffraction (XRD) analysis of bone tissue and coatings was carried out at 2, 4, 12 and 36 weeks after surgery. The crystallographic habit of the implant-facing bone, as well as the structural stability of the coating, were evaluated. For each time period and type of ceramic bone apatite lattice at the interface, no significantly different reference apatite lattice and no foreign peak were recorded. Two weeks after implantation, the bone at the interface was strongly unmineralized in all samples; after 4 weeks, poorly mineralized bone microareas decreased. At 12 weeks, the newly formed bone tissue at the interface with both the new coating and HA coating was shown to be fully mineralized; this crystallographic habit was retained at 36 weeks, when particle release from the tested material was lower compared to the controls. The XRD pattern of bone apatite surrounding the coating particles was unmodified. The innovative coating did not alter the mineralization process at the interface. It improved implant osteointegration, mainly due to a limited release of particles. Consequently, clinical performance of external fixation treatment could be improved by modifying the chemical composition of the implant surface.

Animals↗

PubMed1998

X-ray diffraction of bone at the interface with hydroxyapatite-coated versus uncoated metal implants.

The microstructural characteristics of the newly formed bone tissue at the interface with hydroxyapatite-coated and uncoated stainless steel pins used in an external fracture fixation system have been evaluated. The bone far from the interface was used as a control. Pins were transversally inserted into the diaphyses of sheep tibiae and were loaded in for six weeks. Three sheep received coated pins and two received uncoated pins. Crystallographic habit and mineralization of the implant-facing bone were evaluated. Moreover, lattice parameters of bone apatite were measured and hydroxyapatite (HA) coating degradation was investigated, by means of conventional and microbeam X-ray diffraction (XRD). In coated pins, six weeks after the implantation the newly formed bone tissue at the interface did not reach complete maturation, but the presence of the implant did not alter the apatite lattice structure; the lattice parameters did not show statistically significant variations with respect to those observed in the control bone. In uncoated pins, bone tissue rarely appeared totally mineralized and lattice parameters were significantly different with respect to those observed in the bone far from the implant. HA particles were observed spreading in the bone-facing coated pins; the XRD pattern of bone apatite surrounding HA particles was unmodified. It was concluded that HA coatings improved the bone remodelling process during pin fixation in comparison to uncoated pins and did not alter the crystallographic habit of apatite.

Journal Article↗

PubMed1998

A comparison of hydroxyapatite-coated, titanium-coated, and uncoated tapered external-fixation pins. An in vivo study in sheep.

Three types of surfaces for external fixation pins were compared. One hundred and eight stainless-steel tapered 5/6-millimeter pins were divided into three groups: thirty-six pins remained uncoated (Group A), thirty-six were plasma-sprayed with hydroxyapatite (Group B), and thirty-six were plasma-sprayed with titanium (Group C). The pins were implanted in the left tibia of eighteen sheep, with each sheep receiving six pins from the same group. A unilateral fixator then was assembled on the pins. The medial aspect of the mid-part of the tibial diaphysis was exposed, and a five-millimeter-long cylinder of bone was removed so that load would be borne by the bone-pin interfaces. Six weeks after the procedure, radiographs demonstrated rarefaction of twenty-nine pin tracks in Group A, fifteen in Group B, and thirty in Group C (p = 0.021 for Group A compared with Group B and p = 0.016 for Group B compared with Group C). The mean final insertion torque (and standard deviation) was 4360+/-1050 newton-millimeters in Group A, 3420+/-676 newton-millimeters in Group B, and 3740+/-643 newton-millimeters in Group C. With the numbers available, no significant differences could be detected among these values. The mean extraction torque was 253+/-175 newton-millimeters in Group A, 3360+/-1260 newton-millimeters in Group B, and 1720+/-1030 newton-millimeters in Group C (p = 0.002 for Group A compared with Group B, p = 0.017 for Group A compared with Group C, and p = 0.03 for Group B compared with Group C). The extraction torque was significantly lower than the corresponding insertion torque in both Group A (p < 0.001) and Group C (p = 0.003); no significant difference could be found, with the numbers available, in Group B (hydroxyapatite-coated pins). At sixty times magnification, direct contact was seen along a mean of 16+/-9 per cent of the bone-pin interface in Group A, 30+/-12 per cent of the interface in Group B, and 28+/-15 per cent of the interface in Group C (p = 0.042 for Group A compared with Group C). However, at 10,000 times magnification, direct bone-pin contact was found only in Group B.

Animals↗

PubMed1998

Enhanced fixation with hydroxyapatite coated pins.

Three groups of seven patients had external fixation of middiaphyseal tibial fractures using uncoated pins, uncoated bicylindrical pins, and hydroxyapatite coated bicylindrical pins, respectively. All fractures were fixed with six pins, and all fractures united. Median pin insertion torque was 0.6, 1.2, and 1.3 Nm in the three groups, respectively. Median extraction torque was 0.1, 0.1, and 2.1 Nm, respectively. Both types of stainless steel pins showed a lower extraction torque than insertion torque in all cases, whereas the mean extraction torque in the hydroxyapatite coated pins was unchanged. Seven of the 14 patients receiving uncoated pins had pin tract infection, compared with none of the patients receiving hydroxyapatite coated pins. Hydroxyapatite coating of external fixation pins increases stability and thereby reduces the risk for pin tract infection and mechanical failure of fracture fixation.

Adolescent↗

PubMed1998

Comparison among three different biocoatings for orthopaedic prostheses. An experimental animal study.

One-hundred-eighty cylindrical monocortical titanium implants, 4mm diameter and 12mm long, with three different coatings: fluorohydroxyapatite (group A), hydroxyapatite (group B), and titanium oxide (group C), all applied by vacuum plasma spray were bilaterally, randomly implanted into the femurs and tibiae of twelve adult mongrel sheep. The sheep were divided into four groups (1, 2, 3 and 4) numbering three sheep each. Sheep of groups 1, 2, 3 and 4 were euthanized at two weeks, one month, three and nine months after implantation, respectively Biomechanical and histomorphological analysis were performed. Extraction torque increased over time in all groups until the nine months period. At all the studied periods, the bone-implant contact was higher in Groups A and B compared to Group C. However, only at nine months did this difference reach statistical significance (p<0.005 comparing Groups A and B to C). The results of this study show that all the three coatings could be recommended for clinical applications.

Animals↗

PubMed1998

[Primary cutaneous mucormycosis. A case report].

One case of primary cutaneous mucormycosis developed in the hospital environment is reported. Dressings contaminated by hyphae may have been the cause. In diabetic or immunodepressed patients this disease may be fatal. Incisional biopsy is the only diagnostic tool.

Aged↗

PubMed1997

Surgical treatment of isolated forearm non-union with segmental bone loss.

Twenty-four isolated radius and ulna non-unions with segmental bone loss were operated on. Nine non-unions were in the radius, 15 in the ulna. The surgical technique consisted of removal of the necrotic bone, filling of the bone defect with an intercalary bone graft and internal fixation with a cortical bone graft fixed opposite to a plate. The average length of bone defect after freshening of the bone ends was 3.6 cm (range 2-10.5 cm). The average length of follow-up was 90 months. In 23 cases union was achieved. In three cases a postoperative infection developed. Resolution of the infection after surgical debridement was achieved in two of these patients, while recurrent infection caused failure of the treatment in the third patient. Statistical analysis revealed a shorter healing time (p = 0.031) in the ulna non-unions (12.5 +/- 3.0 weeks) compared with the radius non-unions (16.4 +/- 4.2 weeks). Functional results were classed as excellent in 10 patients, satisfactory in six, unsatisfactory in seven and failure in one.

Adolescent↗