"Full metal jacket" with direct stenting of complete chronic occlusions of the superficial femoral artery.

PURPOSE: This study evaluated the short- and midterm patency of complete total occlusions of the superficial femoral artery (SFA) treated with direct stenting. MATERIALS AND METHODS: Fifty-two consecutive patients (36 men and 16 women; mean age 73.6 years; range 58-85) with chronic complete SFA occlusion and good distal run-off (two or three patent vessels) underwent endovascular recanalisation by direct stenting. All patients were symptomatic (severe claudication or critical ischaemia). Recanalisation was achieved by using a contralateral approach in 44 patients and an ipsilateral antegrade access in eight patients. A total of 152 nitinol stents were used: three stents in 32 cases, four stents in eight cases and two stents in 12 cases. Follow-up consisted of clinical evaluation and colour Doppler ultrasound at 6, 12, 18 and 24 months. RESULTS: The immediate technical success rate was 100%, with complete SFA recanalisation documented on postprocedural angiography. Four cases of distal embolism occurred, which were treated successfully with intra-arterial thrombolysis. During the follow-up, 12 reocclusions were observed: eight were treated with mechanical thrombectomy and in-stent angioplasty, and four were converted into femoropopliteal bypasses. The primary patency rates at 6, 12, 18 and 24 months were 92.3%, 76.9%, 69.2% and 69.2%, respectively. The secondary patency rates at 6, 12, 18 and 24 months were 100%, 100%, 92.3% and 92.3%. CONCLUSIONS: The percutaneous treatment of chronic complete SFA occlusions yielded good primary and secondary patency rates in the short and medium term, with few periprocedural complications. Reocclusions were treated using the percutaneous technique, which guarantees a good secondary patency rate.

Comparative study of jaws with multislice computed tomography and cone-beam computed tomography.

PURPOSE: The aim of this study was to compare the dosimetric and diagnostic performance of multislice computed tomography (MSCT) and cone-beam computed tomography (CBCT) in the study of the dental arches. MATERIALS AND METHODS: Effective dose and dose to the main organs of the head and neck were evaluated by means of thermoluminescent dosimeters (TLDs) placed in an Alderson Rando anthropomorphic phantom and using a standard CBCT protocol and an optimised MSCT protocol. Five patients with occlusal plane ranging from 54 cm to 59 cm who needed close follow-up (range 1-3 months) underwent both examinations. Image quality obtained with CBCT and MSCT was evaluated. RESULTS: Effective dose and dose to the main organs of the head and neck were higher for MSCT than for CBCT. Image quality of CBCT was judged to be equivalent to that of MSCT for visualising teeth and bone but inferior for visualising soft tissues. Beam-hardening artefacts due to dental-care material and implants were weaker at CBCT than at MSCT. CONCLUSIONS: When panoramic radiography is not sufficient in the study of the teeth and jaw bones, CBCT can provide identical information to MSCT, with a considerable dose reduction. MSCT is, however, indicated when evaluation of soft tissue is required.

Percutaneous treatment of complete chronic occlusions of the superficial femoral artery.

PURPOSE: This study was done to evaluate the mid-and long-term patency rates of complete (from the origin to Hunter's duct) chronic occlusions of the superficial femoral artery (SFA) treated by angioplasty and/or stenting. MATERIALS AND METHODS: From February 2002 to March 2005, 21 patients with complete occlusion of the SFA and good distal runoff (two or three patent vessels) were treated with endovascular recanalisation. All patients had severe claudication or critical limb ischaemia. In all cases, recanalisation was performed with a contralateral approach by percutaneous transluminal angioplasty (PTA), with stenting only when PTA provided unsatisfactory results (due to elastic recoil and complications such as dissection). In the case of calcified occlusions and when the true lumen of the SFA could not be crossed, subintimal angioplasty was performed. Follow-up was done at 6 and 12 months and annually thereafter (range 6-55 months, mean 23 months) with clinical evaluation and colour-Doppler ultrasound. RESULTS: Immediate technical success was achieved in all cases (100%), with post-procedural angiography demonstrating complete recanalisation of the SFA. Two distal embolisation (9.5%) occurred, both treated successfully by local thrombolysis. Primary patency rates at 6, 12, 24, 32 and 44 months were 93.3%, 69.2%, 72.7%, 62.5% and 40%, respectively; secondary patency rates at 6, 12 and 24 months were 100%, 84.6% and 81.8%, respectively. CONCLUSIONS: Percutaneous treatment of complete chronic occlusions of the SFA showed good mid-and long-term primary patency rates, with few periprocedural complications. Re-occlusions can be treated by a percutaneous technique, which ensures a good secondary patency rate.

Fluoroscopically guided retrograde replacement of ureteral stents.

PURPOSE: We assessed the feasibility of fluoroscopically guided transurethral replacement of ureteral stents as an alternative to cystoscopy. MATERIALS AND METHODS: Over the last year, we replaced 27 double-J ureteral stents in 20 patients (10 men and 10 women; mean age 67.7 years, range 43-83); 15/20 patients had a native kidney, 3/20 had a transplanted kidney and 2/20 had a ureteroileal conduit. The procedures were performed in the angiography suite with the patient under sedation. All stents were grasped with a gooseneck snare under fluoroscopic control, and the distal end was withdrawn just outside the urethra; then a wire was advanced through the stent lumen and positioned in the renal pelvis. The stent was then removed and replaced with a new double-J stent. RESULTS: The procedures were successful in 26/27 cases. We observed 7 cases of mild haematuria that resolved spontaneously. During follow-up (1-16 months, mean 6.7), stent obstruction occurred in 4 cases, requiring an additional retrograde replacement. CONCLUSIONS: Transurethral fluoroscopically guided retrograde replacement of dysfunctioning ureteral stents is an effective and safe alternative to cystoscopy.

Cloning and analysis of a gene from Streptomyces lividans 66 encoding a novel secreted protease exhibiting homology to subtilisin BPN'.

Amino-terminal degradation has been observed for many of the secreted heterologous proteins produced by S. lividans 66. We, therefore, set out to characterize the relevant proteinases and their genes. A tripeptide chromogenic substrate was used to identify a gene that was shown to encode a secreted protein which removed tripeptides from the amino terminus of extracellular proteins (tripeptidyl aminopeptidase, Tap; Butler et al. 1995). This activity was removed by a homologous gene deletion replacement and the ability of the S. lividans strain to remove N-terminal tripeptides was greatly reduced, but still significant. When the tap-deleted strain was used as a host for the rescreening of a S. lividans 66 genomic DNA library, a number of other genes encoding proteases with aminopeptidase activities were discovered. One clone (P5-4) produced a 45-kDa secreted protein (Ssp), which showed activity against Ala-Pro-Ala-beta-naphthylamide (APA-beta NH-Nap) substrate. Further analysis of the cloned DNA showed an open-reading frame encoding a protein larger than 45 kDa. Direct Edman degradation of the secreted protein confirmed that it was encoded within the cloned DNA and probably processed from a larger precursor. Protein sequence analysis revealed a striking homology to subtilisin BPN' in three regions around the active-site residues suggesting that the protein is a serine protease. As expected, the protease activity was inhibited by phenylmethylsulphonyl fluoride. Mutant strains with most of the ssp gene deleted exhibited reduced activity against APA-beta NH-Nap substrate compared to their non-deleted parental strains.

Isolation and characterization of two genes encoding proteases associated with the mycelium of Streptomyces lividans 66.

A strain of Streptomyces lividans 66 deleted for a major tripeptidyl aminopeptidase (Tap) was used as a host to screen an S. lividans genomic library for clones overexpressing activity against the chromogenic substrate Ala-Pro-Ala-beta-naphthylamide. In addition to reisolation of the tap gene, clones representing another locus, slpD, were uncovered. slpD was analyzed by deletion subcloning to localize its functional sequence. Nucleotide sequence determination revealed an open reading frame encoding a 55-kDa protein exhibiting significant amino acid sequence homology to Tap, particularly around the putative active-site serine residue. No secreted protein was observed for strains harboring the slpD clone, but inspection of the predicted protein sequence revealed a putative lipoprotein signal peptide (signal peptidase II type), suggesting a mycelial location for the SlpD proteinase. In an attempt to isolate an endoprotease known to be active against some heterologous proteins, a second clone was isolated by using a longer substrate (t-butyloxycarbonyl [Boc]-APARSPA-beta-naphthylamide) containing a chemical blocking group at the amino terminus to prevent aminopeptidase cleavage. This locus, slpE, appeared to also encode a 55-kDa mycelium-associated (lipoprotein) proteinase, whose predicted protein sequences showed significant amino acid homology to Tap and SlpD, particularly around the putative active-site serine residues. Chromosomal integration and deletion analysis in both the wild-type and Tap-deficient backgrounds appeared to indicate that SlpD was essential for viability and SlpE was required for growth on minimal media.

Cloning and characterization of a gene encoding a secreted tripeptidyl aminopeptidase from Streptomyces lividans 66.

The gene encoding a tripeptidyl aminopeptidase (Tap) from Streptomyces lividans was cloned by using a simple agar plate activity assay. Overexpression of the cloned gene results in the production of a secreted protein which has an apparent subunit molecular weight of 55,000 and is responsible for the major amino-terminal degradative activity in culture broths of S. lividans strains. A DNA sequence analysis revealed a potential protein-encoding region of the size expected to encode the observed protein, which contained a sequence that exhibited significant homology around a putative active site serine residue observed for lipases, esterases, and acyl transferases. Preceding the amino terminus of the secreted protein was a predicted signal peptide of 36 amino acids followed by a tripeptide, which could be autocatalytically removed from a secreted Tap precursor. The transcriptional start site for the gene was mapped by primer extension. Mutant strains of S. lividans lacking detectable Tap activity were able to grow and sporulate normally. Cross-species hybridization experiments showed that DNA homologs of the tap gene are present in most of the Streptomyces strains tested.

The aminopeptidase N-encoding pepN gene of Streptomyces lividans 66.

The gene (pepN) encoding an aminopeptidase N (PepN) has been cloned from Streptomyces lividans. This was done using either leucine-beta-naphthylamide or arginine-beta-naphthylamide in a liquid overlayer on colonies growing on agar medium to screen for overproduction of the ability to hydrolyse the substrates. The nucleotide sequence of pepN was determined and shown to encode a 95-kDa protein, which displayed significant homology to PepN proteins from other organisms. Analysis of the overproduced proteinase confirmed that this protein was located intracellularly as a monomeric active species. PepN is a metallo-exopeptidase cleaving next to Leu, Arg and Lys in peptide-bond-containing substrates.

Intracellular aminopeptidases in Streptomyces lividans 66.

We have investigated the aminopeptidase activities present in Streptomyces lividans strains. The majority of these activities proved to be intracellular with multiple active species. Two aminopeptidase P genes were identified to be responsible for the ability to hydrolyze amino terminal peptide bonds adjacent to proline residues. Two other broad spectrum aminopeptidases were found to display homology at both the DNA and protein levels. One showed significant homology to PepN proteins, particularly around the putative zinc-binding residues which are important for catalysis. The second broad spectrum activity was not analyzed in detail but showed a different spectrum of substrate specificity to that of PepN.