An intervention program for university students who have parents with alcohol problems: a randomized controlled trial.

AIM: To study the effects of alcohol and coping intervention among University students who have parents with alcohol problems. METHODS: A total of 82 university students (56 women and 22 men, average age 25) with at least one parent with alcohol problems were included. The students were randomly assigned to one of three programs: (i) alcohol intervention program, (ii) coping intervention program, and (iii) combination program. All programs were manual based and individually implemented during two 2-h sessions, 4 weeks apart. This assessment contained both a face-to-face interview and six self-completion questionnaires; AUDIT, SIP, EBAC, coping with parents' abuse questionnaire, SCL-90 and ISSI. Follow-up interviews were conducted after 1 year. RESULTS: All participants finished the baseline assessment, accepted and completed the intervention, while 95% of the students completed the 12-month follow-up assessment. The two groups that received alcohol intervention improved their drinking pattern significantly more than the group that did not receive alcohol intervention [change of standardized scores -0.27 (CI -0.53 to -0.03)]. The groups receiving coping intervention did not differ from the group not receiving coping intervention concerning their ability to cope with their parents' alcohol problems. Nor did they differ regarding changes in their own mental health or in their social interaction capacity. CONCLUSION: The intervention improved drinking patterns in adult children of alcoholics.

A combined RSA and FE study of the implanted proximal tibia: correlation of the post-operative mechanical environment with implant migration.

There is strong evidence to suggest that inducible displacements, migration and implant loosening are closely related to the initial mechanical environment of the implanted tibia. If this is true, then it should be possible to predict the likelihood of implant migration using patient-specific finite element models. Finite element models of the proximal implanted tibiae were analysed based on pre-operative quantitative computed tomography data of four patients entered into a radiographic migration study. These four patients were also part of an radiostereometric analysis (RSA) study. A variety of load cases were analysed and the risk of bone failure determined for a 2 mm layer of bone immediately beneath the tibial tray. The results were compared with the RSA data measured 1 year post-operatively for each patient. For each patient, an appropriate load case was selected based on patient weight and on the varus-valgus migrations observed in the migration study. The two patients with press-fit implants were predicted to have the highest risk of failure and were found to migrate the most. The two patients with bonded implants (one HA coated and one cemented) were found to have a low risk of failure and these implants migrated the least. This study suggests that the degree of implant migration is dependent on the initial mechanical environment and can be determined using patient-specific finite element analysis.

Inter- and intramolecular domain interactions of the catalase-peroxidase KatG from M. tuberculosis.

The inter- and intramolecular interactions between the different domains of the catalase-peroxidase KatG from Mycobacterium tuberculosis were analyzed using the two-hybrid assay. It was shown that the dimerization of the enzyme is due to a strong interaction of the first 99 amino acids of the N-terminal domain whereas the C-terminal domain does not play a role in the dimerization. In addition, an intramolecular interaction between the N- and C-terminal domains was detected which might play a functional role in the mechanism of the enzyme.

Changing the substrate specificity of cytochrome c peroxidase using directed evolution.

Cytochrome c peroxidase (CCP) from Saccharomyces cerevisiae was subjected to directed molecular evolution to generate mutants with increased activity against 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). Using a combination of DNA shuffling and saturation mutagenesis, mutants were isolated which possessed more than 20-fold increased activity against ABTS and a 70-fold increased specificity toward ABTS compared to the natural substrate. In contrast, activities against another small organic molecule, guaiacol, were not significantly affected. Mutations at residues Asp224 and Asp217 were responsible for this increase in activity. These two residues are located on the surface of the protein and not in the direct vicinity of the distal cavity of the peroxidase, where small organic substrates are believed to be oxidized. Mutations at position Asp224 also lead to an increased amount of the active holoenzyme expressed in Escherichia coli, favoring the selection of these mutants in the employed colony screen. Possible explanations for the effect of the mutations on the in vitro activity of CCP as well as the increased amount of holoenzyme are discussed.

Directed molecular evolution of cytochrome c peroxidase.

Cytochrome c peroxidase (CCP) from Saccharomyces cerevisiae was subjected to directed molecular evolution to generate mutants with increased activity against the classical peroxidase substrate guaiacol, thus changing the substrate specificity of CCP from the protein cytochrome c to a small organic molecule. After three rounds of DNA shuffling and screening, mutants were isolated which possessed a 300-fold increased activity against guaiacol and an up to 1000-fold increased specificity for this substrate relative to that for the natural substrate. In all of the selected mutants, the distal arginine (Arg48), which is fully conserved in the superfamily of peroxidases, was mutated to histidine, showing that this mutation plays a key role in the significant increase in activity against phenolic substrates. The results suggest that, in addition to stabilizing the reactive intermediate compound I, the distal arginine plays an important role as a gatekeeper in the active site of CCP, controlling the access to the ferryl oxygen and the distal histidine. Other isolated mutations increase the general reactivity of the peroxidase or increase the intracellular concentration of the active holo form, allowing their selection under the employed screening conditions. The results illustrate the ability of directed molecular evolution technologies to deliver solutions to biochemical problems that would not be readily predicted by rational design.

Spontaneous Formation of the Bioactive Form of the Tuberculosis Drug Isoniazid.

It works even without the enzyme: The formation of the bioactive form 1 of isoniazid inside Mycobacterium tuberculosis does not depend on specific interactions within the active site of the inhibited enzyme, but rather relies on the fast addition of acyl radicals to electron-deficient heterocycles such as NAD(+).

Use of site-directed mutagenesis to probe the structure, function and isoniazid activation of the catalase/peroxidase, KatG, from Mycobacterium tuberculosis.

A series of mutants bearing single amino acid substitutions often encountered in the catalase/peroxidase, KatG, from isoniazid-resistant isolates of Mycobacterium tuberculosis has been produced by site-directed mutagenesis. The resultant enzymes were overexpressed, purified and characterized. Replacing Cys-20 by Ser abolished disulphide-bridge formation, but did not affect either dimerization of the enzyme or catalysis. The substitution of Thr-275, which is probably involved in electron transfer from the haem, by proline resulted in a highly unstable enzyme with insignificant enzyme activities. The most commonly occurring substitution in drug-resistant clinical isolates is the replacement of Ser-315 by Thr; this lowered catalase and peroxidase activities by 50% and caused a significant decrease in the KatG-mediated inhibition of the activity of the NADH-dependent enoyl-[acyl-carrier protein] reductase, InhA, in vitro. The ability of this enzyme to produce free radicals from isoniazid was severely impaired, as judged by its loss of NitroBlue Tetrazolium reduction activity. Replacement of Leu-587 by Pro resulted in marked instability of KatG, indicating that the C-terminal domain is also important for structural and functional integrity.

Overexpression, purification, and characterization of the catalase-peroxidase KatG from Mycobacterium tuberculosis.

Wild-type catalase-peroxidase KatG from Mycobacterium tuberculosis as well as a specific mutant (R463L) frequently found in isoniazid-resistant strains have been overexpressed in Escherichia coli, allowing purification of sufficient quantities of enzyme for physical and kinetic characterization. Optical absorption and EPR spectroscopies indicate that KatG is similar to a growing class of bacterial catalase-peroxidases. Optical and EPR spectra of KatG in the presence of either a strong field or weak field ligand suggest that, like horseradish peroxidase and metmyoglobin, KatG is likely to have a histidine as a proximal ligand. The wild-type enzyme functions as a highly active catalase as well as a broad specificity peroxidase. Wild-type KatG and the R463L mutant of KatG exhibit identical spectroscopic and kinetic properties. Furthermore, both enzymes are equally capable of metabolizing the important antituberculosis drug isoniazid.

Organization of 'nanocrystal molecules' using DNA.

Patterning matter on the nanometre scale is an important objective of current materials chemistry and physics. It is driven by both the need to further miniaturize electronic components and the fact that at the nanometre scale, materials properties are strongly size-dependent and thus can be tuned sensitively. In nanoscale crystals, quantum size effects and the large number of surface atoms influence the, chemical, electronic, magnetic and optical behaviour. 'Top-down' (for example, lithographic) methods for nanoscale manipulation reach only to the upper end of the nanometre regime; but whereas 'bottom-up' wet chemical techniques allow for the preparation of mono-disperse, defect-free crystallites just 1-10 nm in size, ways to control the structure of nanocrystal assemblies are scarce. Here we describe a strategy for the synthesis of 'nanocrystal molecules', in which discrete numbers of gold nanocrystals are organized into spatially defined structures based on Watson-Crick base-pairing interactions. We attach single-stranded DNA oligonucleotides of defined length and sequence to individual nanocrystals, and these assemble into dimers and trimers on addition of a complementary single-stranded DNA template. We anticipate that this approach should allow the construction of more complex two- and three-dimensional assemblies.

Lumbar spinal stenosis. A retrospective study of 163 cases in southern Sweden.

I assessed the incidence of lumbar spinal stenosis as well as the frequency of severe neurological symptoms and signs based on patients seen in 2 orthopedic departments. The annual incidence in Malmõ, 1982-1986, was 59 and in the period 1987-1991, 47 per million inhabitants. In Växjö, 1987-1991, the annual incidence was 45 per million. Severe neurological symptoms were few. Reduced EHL power and peroneal paresis were the most prevalent signs and were found in 13 and 12 percent, respectively, of all 163 patients. 1 patient had bladder dysfunction, 1 had impotence and 2 had a cauda equina syndrome.

Measuring hallux valgus: a comparison of conventional radiography and clinical parameters with regard to measurement accuracy.

To assess the repeatability and error of conventional x-ray measurements, intra- and interobserver evaluations of measurement accuracy were done on 20 preoperative and 40 postoperative (20 chevron and 20 proximal osteotomy) x-rays of hallux valgus patients. Standard x-rays showed an average interobserver error of measurement of 6.4 degrees for the hallux valgus angle, 5.4 degrees for the intermetatarsal angle, and 2.0 mm for the intermetatarsal distance. The intraobserver error did not differ greatly. The repeatability and error of two clinical measurements, ball circumference, and dorsal to plantar range of motion of the first metatarsophalangeal joint were evaluated for 20 healthy volunteers. The ball circumference had an average measurement error of 1.1 cm, whereas the dorsal and plantar range of motion of the great toe had an average measurement error of 12 degrees in dorsiflexion and 16 degrees in plantarflexion. In both clinical and radiographic parameters, linear measurements were more accurate than angular measurements. Although x-rays are of value in hallux valgus surgery, standard x-rays are less accurate than previously assumed. Small changes produced by osteotomies may be hidden by the postoperative measurement error. The results of hallux valgus surgery should primarily be evaluated clinically. When clinical and radiological evaluations are made, linear measurements may be preferable.

Hip involvement in early rheumatoid arthritis.

OBJECTIVE: To study early hip involvement in rheumatoid arthritis (RA) and to evaluate the usefulness of ultrasonography in the detection of hip joint synovitis in RA. METHODS: Study I: The number of hip joint replacements was recorded in a cohort of 113 patients with RA of at least five years disease duration followed from an early stage. Study II: Ultrasonography was evaluated as a method to identify hip joint synovitis in 76 patients with RA of shorter disease duration, by relating it to radiograms and clinical findings. RESULTS: Study I: Twenty one hip joint replacements were performed in 15 of the 113 patients. The median disease duration at the time of first arthroplasty was 48 (range 10-76) months; the annual incidence was approximately constant between two and six years. High disease activity at the start of the study was predictive of requirement for hip prosthesis. Study II: Hip ultrasonography was pathological in 13 of the 76 patients studied, bilaterally in nine. Hip joint synovitis could not be confirmed on clinical grounds only as seven of the patients with positive ultrasonographic findings were asymptomatic, and the remaining six patients had only mild symptoms of hip involvement. Also, six of the 63 patients with normal ultrasonography had mild symptoms. There was no difference regarding demographic, clinical, and laboratory findings in patients with and without hip synovitis. CONCLUSIONS: Early hip joint destruction giving symptoms mostly at a very late stage is frequent in RA. Ultrasonography rather than signs or symptoms could identify patients with hip joint involvement and provide a rationale for early treatment.

Synthesis, structure and activity of artificial, rationally designed catalytic polypeptides.

Biological macromolecules with catalytic activity can be created artificially using two approaches. The first exploits a system that selects a few catalytically active biomolecules from a large pool of randomly generated (and largely inactive) molecules. Catalytic antibodies and many catalytic RNA molecules are obtained in this way. The second involves rational design of a biomolecule that folds in solution to present to the substrate an array of catalytic functional groups. Here we report the synthesis of rationally designed polypeptides that catalyse the decarboxylation of oxaloacetate via an imine intermediate. We determine the secondary structures of the polypeptides by two-dimensional NMR spectroscopy. We are able to trap and identify intermediates in the catalytic cycle, and to explore the kinetics in detail. The formation of the imine by our artificial oxaloacetate decarboxylases is three to four orders of magnitude faster than can be achieved with simple amine catalysts: this performance rivals that of typical catalytic antibodies.

The effects of hyperbaric oxygenation on bone-titanium implant interface strength with and without preceding irradiation.

This study investigated the influence of a single 15-Gy dose of irradiation on the capacity of titanium screws to integrate in irradiated bone tissue. The biomechanical force necessary to unscrew the titanium implants 8 weeks after placement was 54% lower for implants in irradiated bone tissue compared to implants in nonirradiated bone tissue. Postirradiation use of hyperbaric oxygen treatment at 2.8 ATA (2-hour daily treatments for 21 days) increased the biomechanical force necessary to unscrew the titanium implants by 44% in irradiated bone and by 22% in nonirradiated bone.

The natural course of lumbar spinal stenosis.

The course of 32 untreated patients with spinal stenosis was studied. The mean patient age was 60 years, and the mean period of observation was 49 months. About 75% of the patients had spinal claudication. In the follow-up survey, the same number of patients had claudication, but the symptoms were milder. In estimation by visual analog scale, symptoms in 70% of the cases were unchanged, 15% showed improvement, and 15% worsened. No proof of severe deterioration was found after four years, and expectant observation may be an alternative to surgical treatment.

The effect of decompression on the natural course of spinal stenosis. A comparison of surgically treated and untreated patients.

The clinical course of 19 untreated patients with spinal stenosis (mean age, 60 years) was compared with that of 44 patients treated surgically (mean age, 65 years). The time of follow-up was 31 and 53 months, respectively. About 80% of the patients had neurogenic intermittent claudication. In the follow-up, one third of the treated and one half of the untreated patients still had neurogenic claudication. By visual analogue-scale estimation, 60% of those treated surgically and 33% of the untreated patients felt better. Fifty-eight percent of the untreated patients were unchanged. Neurophysiologic changes showed progression in almost all cases; it was more pronounced in the treated patients. No proof of severe deterioration was found in the untreated patients, and observation for 2-3 years seems to be a good alternative to surgery.