Upper body lift and breast reshaping with lateral chest wall perforator propeller flap following massive weight loss.

BACKGROUND: After massive weight loss (MWL), female patients often develop upper trunk laxity and severe breast deformities. Usually several procedures are required to address upper body contouring issues. OBJECTIVES: To achieve better breasts and improve upper body contour, the authors employed a combined approach, associating lateral chest wall perforator propeller flaps with an upper bodylift (UBL). METHODS: Between September 2015 and March 2017, nine post-bariatric patients underwent simultaneously an UBL and autologous augmentation breast reshaping with lateral chest wall perforator propeller flaps. The authors analyzed the clinical indications, results and complications of this procedure. RESULTS: Eighteen lateral perforator propeller flaps for autologous breast augmentation-mastopexy associated with an UBL were performed successfully. Mean pre-MWL body mass index (BMI) was 54.3±10.9kg/m2, with a mean preoperative pre-UBL BMI of 28.7±3.6kg/m2. The average weight loss before surgery was 67.7±22.4kg. The flaps were harvested on intercostal and/or lateral thoracic arteries. All donor sites had been closed primarily. Following the classification of Dindo and Clavien, four minor complications (I, II), and two major complications (IIIb), including two hematomas requiring reoperation, were reported. No flap necrosis occurred. Follow-up averaged 27.9±8.4months. The patients' satisfaction with their improved breast shapes and chest wall contours was "good", with an aesthetic outcome mean ranked 3.8±0.8 (out of 5). CONCLUSIONS: After MWL, upper body deformities can be treated safely and reliably by a combined approach, associating an UBL and autologous lateral chest wall perforator flaps to provide more natural and durable breast shapes, as well as an upper circumferential reshaping.

Thoracotomy and esophageal surgery: Key points to preserve the possibilities of flaps.

Anastomotic leakage frequently complicates esophagectomy and can trigger a rare life- threatening complication, a tracheoesophageal fistula. No guideline has yet addressed this complication. Plastic surgeons play a crucial role for salvage surgery. When a re-operation is chosen the possibilities of flap interposition depend on how the thoracotomy was initially performed. This study tried to identify key techniques in order help thoracic or general surgeons to preserve all the local flaps available for TEF if it occurs. These techniques improve flap conservation, helping plastic surgeons when a later transposition flap is required.

[Fournier's gangrene: Cervical and facial extension. A very rare case]. / Gangrène gazeuse périnéo-fessière à extension cervico-faciale. À propos d'un cas exceptionnel.

Fournier's gangrene is a fearsome disease with a bad prognosis and a mortality rate ranging between 10 and 80% according to the literature. It is extensive in 13 to 54% of cases. Up to date, cervico-facial extension has never been reported. We describe the case of a 51-year-old overweighed woman with a history of type 2 diabetes and a narrow lumbar canal who was referred to our institution for significant fatigue and increasingly painful legs. A diagnosis of Fournier's gangrene was made after correlating the physical findings with the results of a full body scan. Diffuse subcutaneous emphysema involving the face, neck, mediastinum, abdominal wall, right buttock, perineum and the right thigh was identified. Treatment included multiple surgical debridements, admission to intensive care unit, and an efficient antibiotic therapy that enabled preservation of the patient's life. To our knowledge, this is the first case of cervical and mediastinal extension of Fournier's gangrene to be reported. No clear guidelines exit on the management of this complication (cervico-facial and mediastinal drainage). We share our experience of this unusual case.

Bovine corneal protein 54K (BCP54) is a homologue of the tumor-associated (class 3) rat aldehyde dehydrogenase (RATALD).

Amino acid (aa) sequence data from Staphylococcus areas V8 protease-digested bovine corneal 54-kDa protein (BCP54) fragments were utilized to derive mixed oligodeoxyribonucleotide (oligo) primers complementary to the reverse translation products of these sequences. These degenerate oligo primers were used to prime the amplification of BCP54 sequence from bovine corneal epithelial cell cDNA. The cDNA probe generated by this mixed oligo-primed amplification of cDNA was cloned and dideoxy-sequenced. A search of the GenBank database (version 63.0) revealed extensive sequence similarity to the cDNA encoding tumor-associated rat liver (class 3) aldehyde dehydrogenase (RATALD). Nucleotide (nt) and aa sequence alignment of the BCP54 translation product reveals it is 78% and 84% homologous with RATALD at the nt and aa levels, respectively. Conservation of aa sequence elements common to the aldehyde dehydrogenase family thought to be of structural/functional significance is further substantiated by this analysis. Included in the discussion is the likelihood that gene sharing (genes encoding metabolic enzymes and other stable proteins) may extend to the cornea.

Full-length cDNA cloning utilizing the polymerase chain reaction, a degenerate oligonucleotide sequence and a universal mRNA primer.

A full-length cDNA was selectively amplified by the polymerase chain reaction (PCR) utilizing a primer pair consisting of a "universal" 21-base synthetic deoxyoligonucleotide (oligo dT 17GGCC) and a specific degenerate deoxyoligonucleotide sequence (DOS) derived from the N-terminal amino acid sequence. This double-stranded amplified cDNA was uni-directionally cloned into M13mp19 utilizing two restriction sites that had been previously incorporated into the termini of the universal and specific DOS primers. Cloning of the specific cDNA via this PCR amplification with the universal/specific DOS primer pair approach was confirmed by screening with a second DOS contiguous with the DOS employed to prime second (sense)-strand cDNA synthesis. This technique allows for the selective full-length cDNA cloning of low-abundance mRNAs from a single-protein sequence determination.

Screening recombinant DNA libraries: a rapid and efficient method for isolating cDNA clones utilizing the PCR.

We describe an expeditious method for the isolation of cDNA clones utilizing PCR-based amplification of target sequences from cDNA libraries. This method is rapid, less labor-intensive and inexpensive when compared with screening libraries with radiolabeled probes. This method can be applied to isolate multiple members of a protein family as well as homologous genes in different species by designing appropriate primers to amplify the most conserved regions. Utilizing this method, a novel reticulocyte CD44 transcript was isolated.

Laser desorption mass spectrometry for microbial DNA analysis.

Recently, we demonstrated that a matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI-TOF MS) can be used to determine the molecular weight of polymerase chain reaction (PCR) products of intact 16S rRNA regions and to profile their restriction digests. This is the first time that MALDI-TOF MS with ultraviolet (UV) photoionization has been used to analyze a PCR product of approximately 1600 nucleotides in length.

PCR-Based Full-Length cDNA Cloning Utilizing the Universal-Adaptor/Specific DOS Primer-Pair Strategy.

Biological systems are often influenced by molecules that are neither present in vast quantities or easily purified to homogeneity from other cellular constituents. The development of simple, efficient molecular cloning systems coupled with the relative ease of DNA sequence determination has made nucleic acid sequence determination the choice methodology when sequence determination of the complete biological molecule is indicated. However, one bottleneck that impedes direct access to sequence determination is the necessity to screen recombinant libraries containing a large number of clones for the low-abundance member. Advances in protein microsequencing techniques combined with application of the polymerase chain reaction (PCR) help simplify this process (1-3). Utilizing only the N-terminal protein sequence information, we have developed a protocol for the selective amplification and subsequent cloning of specific full-length cDNAs, which in some instances may circumvent the time-consuming, tedious process of library screening. This approach, as illustrated in Fig. 1, employs the following steps:

Discrimination between tumour and normal cells by staining with 3,4,5,6,16,17-hexadehydro-16-(methoxycarbonyl)-19 alpha-methyl-20 alpha-oxayohimbanium: the uracil ring as a target for the specific interaction between RNA(s) and the fluorescent probe.

3,4,5,6,16,17-Hexadehydro-16-(methoxycarbonyl)-19 alpha-methyl-20 alpha-oxayohimbanium (Alstonine) is a fluorescent alcaloid which has been known to stain tumour cells more efficiently than normal ones. In this paper the spectral properties of Alstonine were first investigated and its capability for preferential staining of tumour cells verified in culture using SK-OV-3 cells as tumour cells and Mouse 3T3 fibroblasts as controls. Then interactions between Alstonine and biological macromolecules were investigated to provide the rationale for preferential labelling. Molecular filtration techniques have demonstrated that binding occurs only with RNA molecules. Similar experiments were performed with different isopolynucleotides to find an explanation for that specificity. They provide evidence that binding occurs only in the presence of a uridyl ring. This is consistent with the specificity of the linkage to RNA. As the linkage of Alstonine with RNA did not induce any shift or obvious change in the intensity of its fluorescence spectrum, it is concluded that the binding might involve the side chain of the fluorescent compound.

Partial amino acid sequence determination of bovine corneal protein 54 K (BCP 54)

The most abundant soluble protein of bovine cornea, BCP 54 (Bovine Corneal Protein, molecular weight 54 kD) was isolated and digested under both limited and complete digestion conditions with Staphylococcus aureus V8 protease. The fragments resulting from limited digestion were separated by one-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis, transferred to a polyvinylidene difluoride membrane, visualized by Coomassie Blue staining, cut out, and submitted to N-terminal protein sequence analysis. Complete digestion fragments were separated on a Vydac C18 reverse-phase HPLC, collected, and concentrated prior to sequencing. Using this method, we obtained amino acid sequence data from three internal V8 protease derived fragments of BCP 54 and a number of HPLC fragments. Comparison of these amino acid sequences, corresponding to 30% of the BCP 54 molecule, to those sequences contained within release 22 of the National Biomedical Research Foundation Protein Identification Resource revealed no extended sequence similarity of known proteins to BCP54.

Biotin-enhanced fragmentation for direct deoxyribonucleic acid sequencing using matrix-assisted laser desorption/ionization mass spectrometry.

Fragmentation of synthetic oligonucleotides under the influence of biotin was investigated using 3-hydroxypicolinic acid (3-HPA) as a matrix-assisted laser desorption/ionization (MALDI) matrix. Addition of biotin into the sample enhanced fragmentation of the oligonucleotide between bases. However, when the biotin was tagged to the 5'-terminus of the oligonucleotide, enhancements were observed not only in desorption/ionization efficiency but also in the fragmentation of molecular ions. The protonation/deprotonation process occurs on the tagged biotin is a possible reason for the enhancement in desorption/ionization. Site-specific backbone cleavage fragmentation patterns were observed. The sequences of oligonucleotides can be obtained from their fragment ions. The direct sequencing of a 5'-biotin-tagged 25-mer is demonstrated.

Members of the ALDH gene family are lens and corneal crystallins.

Many of the major lens proteins, known as crystallins, responsible for the structural integrity and functional utility of this visual tissue have been previously shown to be recruited proteins. This phenomena of a protein that is expressed and functions elsewhere acquiring a new function in another tissue has been termed 'gene sharing'. It is now becoming obvious that the cornea of vertebrates has similarly acquired proteins, and that at least one corneal protein, ALDH3 belongs to a gene family that has been previously identified as a lens crystallin. The recognition that both lens and corneal crystallins exist is a novel concept that has implications that involve the process by which multifunctional gene products have evolved. Members of the ALDH gene family function in both the cornea and lens as crystallins and the acquisition of multifunctionality by this gene family is unique. Based on our analysis we have deduced a supragene family relationship between the thiol protein esterases, aldehyde dehydrogenases, and the taxon-specific crystallins. Evolution of a complex organ such as the vertebrate eye is not a sequential and gradual process such as the Darwinian Giraffe's neck, since the eye can provide selective advantage only as a complete organ. Catastrophic theory proposes that the complex vertebrate eye with its lens, and focussing mechanism arose from the primitive eye spot which contained originally only the photoreceptor system by a one step event. In the evolution of the vertebrate eye it is evolutionarily plausible that several pre-existing proteins have been recruited to perform a structural role for this complex organ. It is also incumbent in evolutionary thought that any inherent enzymatic activity associated with this protein would be purely an incidental addition to the organ. However, the fact that most of these have pyridine nucleotide binding capacity, which is presumed important in giving protection from UV exposure, is noteworthy. Finally, to construct the vertebrate eye in one step from the existing visual pigment system such as the eyespot of unicellular organisms the following criteria would apparently be advantageous: (1) high water solubility; (2) transparency; and (3) common genetic regulatory elements (e.g. promoters/enhancers). Although it is an important observation that certain members of the aldehyde dehydrogenase gene family are present as structural proteins in the cornea and lens, it is not surprising that the phenomenon of gene sharing extends to another ocular tissue such as the cornea. In this context, it will be interesting to note if similar multifunctional gene products will be found as frequently in organs other than the eye.

Synthesis and characterization of SERS gene probe for BRCA-1 (breast cancer).

A protocol for binding cresyl fast violet (CFV), a SERS-active dye (label) containing an aromatic amino group with a modified oligomer having a carboxy derivatized thymidine moiety using carbodiimide coupling has been achieved for the first time. Covalent coupling between CFV and the oligomer has been confirmed by mass spectral analysis of the labeled oligomer. The fluorescence, SERS and absorption characteristics of the labeled product have been evaluated. The chosen oligomer contains a BRCA-1 (breast cancer) sequence, and hence has the potential for being used as a gene probe to identify BRCA-1 gene. It has high potential for being used in polymerase chain reaction (PCR) amplification, as has been performed with labeled oligonucleotide for the HIV sequence.

Surface-enhanced Raman gene probe for HIV detection.

We report, for the first time, the use of surface-enhanced Raman (SERS)-active labels for primers used in polymerase chain reaction amplification of specific target DNA sequences. This method has the potential for combining the spectral selectivity and high sensitivity of the SERS technique with the inherent molecular specificity offered by DNA sequence hybridization. The effectiveness of the detection scheme is demonstrated using the gag gene sequence of the human immunodeficiency virus. The potential use of multiple probes for simultaneous detection of multiple biological targets is discussed.

The multispecific cell adhesion molecule CD44 is represented in reticulocyte cDNA.

Identified originally as erythrocyte p80, whose expression is down regulated by the Lutheran inhibitor gene (In[Lu]), the In(Lu) related-p80 glycoprotein represents the red cell isoform of the human cell adhesion/recognition molecule CD44. The presence of a CD44 transcript within a reticulocyte cDNA library was indicated by the PCR amplification of an appropriately sized product generated by a pair of deoxyoligonucleotide primers derived from CD44 cDNA sequence. The amplified product was subsequently utilized to screen and isolate a positively hybridizing full-length reticulocyte cDNA clone (RETIC CD44) that contained an 1809 base pair insert that was DNA sequenced by the dideoxy chain termination method. This first isolate of a reticulocyte CD44 cDNA appears to be generated by a combination of RNA processing events that includes production of 3' mRNA heterogeneity by utilization of multiple poly(A) sites. Remarkably, the 3' untranslated (3'UT) region of this mRNA, encoding a prototypic hematopoietic CD44 isoform, has been previously reported present on only a transcript that encodes a CD44 epithelial isoform.

Culture of Mycobacterium tuberculosis from non-fixed sputum smears.

Smears prepared from sputum species were kept in the dark at 4 degrees C or at 20-25 degrees C. Cultures for mycobacteria were carried out on these smears 0, 4, 8, and 15 days later and the results compared with those of cultures made from the sputum specimen on the day of collection. The percentage of negative cultures from originally positive specimens was high, and use of the method is not considered advisable. There is an additional risk of infection for laboratory staff when handling unfixed smears.

Mycobacteria in HIV-infected patients in Buenos Aires.

SETTING: F. J. Muñiz Hospital and Department of Phthisiopneumonology, in Buenos Aires. OBJECTIVE: To analyze bacteriological findings concerning tuberculosis and other mycobacteriosis, in association with HIV infection and AIDS. DESIGN: From June 1985 to December 1991, 2521 samples from 1259 HIV-seropositive and AIDS patients were analyzed: 1133 samples were of bronchopulmonary origin and the remaining 1388 of extrapulmonary origin. Drug susceptibility tests were performed using the proportions method. RESULTS: Mycobacterial disease was confirmed by culture in 240 of the 1259 HIV/AIDS patients (19%). Mycobacterium tuberculosis was isolated in 223 of these cases (92.9%) and M. bovis in two, while M. avium-complex (MAC) strains were identified as the cause of disease in 14 patients (5.8%). In only one case was disease due to M. kansasii. Blood cultures were positive in 21.2% of these 240 cases. Resistance of M. tuberculosis to antituberculosis drugs was found in 9.4% of the 223 isolates. In only one case was multidrug resistance detected, in a patient who had received previous treatment. CONCLUSION: Smear examination, although less sensitive than in HIV-negative patients, was still a simple and reliable tool for the rapid diagnosis of mycobacterial disease. Blood culture aided in the successful diagnosis of about half of the cases of disseminated tuberculosis and of all cases of MAC disease. An alarming spread of tuberculosis was detected among a group of HIV-positive prisoners, and the possible emergence of multidrug resistance should be anticipated.

PIP: An increase in human immunodeficiency virus (HIV)-associated mycobacterial tuberculosis has led to a reversal of an earlier trend in Argentina toward a decline in the incidence of tuberculosis. A bacteriologic study conducted at the Muniz Hospital in Buenos Aires June 1985-December 1991 confirmed the reliability of smear examination for the rapid diagnosis of mycobacterial disease. 2521 samples were obtained from 1259 HIV-infected individuals during the study period and mycobacterial disease was confirmed by culture in 240 cases (19%). The smears were positive in 59.0% of the 122 pulmonary cases, 22.0% of the 72 extrapulmonary cases, and 56.5% of the 46 pulmonary-extrapulmonary cases. Blood cultures were positive in 21.2% of the 240 cases. In patients with pulmonary localization, acquired immunodeficiency syndrome (AIDS) was diagnosed between a few months to two years after the onset of tuberculosis; those with the two other localizations had already advanced to AIDS at the time of blood smear. Resistance to one or more antitubercular drugs was found in 9.4% of cases.

Chemical cleavage sequencing of DNA using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

In this paper, we report for the first time use of laser desorption mass spectrometry for measurement of chemical cleavage sequencing products of DNA. In this method, the target DNA was labeled with biotin and subjected to chemical modification and cleavage according to the Maxam-Gilbert sequencing protocol. The biotin-containing fragments were captured by streptavidin-coated magnetic beads and separated from the other fragments. The captured fragments were released by hot ammonia treatment, and the released fragments were analyzed by mass spectrometry. Potential applications of this method in resolving sequence ambiguities and sequencing repeat sequences as well as in the analysis of DNA-protein interactions are discussed.