Metabolic syndrome in adolescents in the Balearic Islands, a Mediterranean region.

BACKGROUND AND AIMS: To determine the prevalence of metabolic syndrome (MetS) and its components in adolescents in the Balearic Islands, in the western Mediterranean Sea. METHODS AND RESULTS: A cross-sectional nutritional survey was carried out in the Balearic Islands (2007-2008). A random sample (n=362, 143 boys and 219 girls) of the adolescent population (12-17 years) was interviewed, anthropometrically measured, and provided a fasting blood sample. The MetS prevalence was determined by the ATP III criteria adapted for youths. Adherence to the Mediterranean diet (MD) was defined according to a score constructed considering the consumption of MD components: high monounsaturated fatty acids (MUFA)/saturated fatty acids (SFA) ratio, moderate ethanol consumption, high legume, cereals and roots, fruit, vegetables, and fish consumption, and low meat and milk consumption. The overall MetS prevalence was 5.8% (boys 10.5%, girls 2.7%). MetS criteria were met by 10.0% of overweight, 45.5% of obese and in 1.8% of normal weight adolescents. Half of the adolescents (49.7%) had at least one MetS component. None of the adolescents had all five risk factors. High triglyceride level (90.5%), hypertension (85.7%), low HDL cholesterol level (78.9%) and central obesity (71.4%) were common among adolescents with MetS whereas hyperglycaemia (0.6%) was infrequent. Higher adherence to MD was associated with significantly lower odds ratio of having MetS, but half of the adolescents showed high adherence to MD. CONCLUSION: MetS prevalence was significant among adolescents in the Balearic Islands, especially among obese boys. A high adherence to MD in adolescents was associated with a low prevalence of the MetS criteria.

A rapid and sensitive cellular enzyme-linked immunoabsorbent assay (CELISA) for the detection and quantitation of antibodies against cell surface determinants. I. A comparison of cell fixation and storage techniques.

A solid phase cellular ELISA was designed and evaluated for the detection of antibodies specific for cell surface determinants. It was hypothesized that certain fixation and freezing procedures would result in stabilization of cell structures for prevention of antigen diffusion and extraction during washing procedures. This would assure assay accuracy and convenient sample management. It was hypothesized that fixation with certain reagents prior to analysis would not alter antigenicity of antibody targeted epitopes. In order to improve the preservation of the cells following cell binding to the solid phase matrix while still retaining antigenicity and morphology, a series of fixatives and storage procedures were screened to determine which were best suited for CELISA. Methanol, washing buffer (WB), Hanks' balanced salt solution (HBSS), and 0.5% formalin in HBSS were examined by comparing their relative cell binding capacity and the subsequent cell morphology. In consideration of all variables, fixation in 0.5% formalin provided the best maintenance of cell antigenicity, morphology, binding, and was associated with consistent results. Cells used immediately after fixation and fixed cells used after storage at -80 degrees C for up to 12 months were compared to determine if long term storage affected antigenicity. Since frozen cells and fresh cells demonstrated statistically identical positive to negative ratios and consistency of antibody binding, it was determined that long term frozen storage of formalin-fixed cells did not adversely affect antibody binding capacity to cell surface determinants.

A rapid and sensitive cellular enzyme-linked immunoabsorbent assay (CELISA) for the detection and quantitation of antibodies against cell surface determinants. II. Optimal reagent concentrations and predictive analysis.

A cellular enzyme-linked immunosorbent assay (CELISA) was developed for the detection and quantification of antibodies elicited against allogeneic cell surface determinants. The technique uses a solid-phase cell matrix created by fixing cells with a mild formalin solution onto the bottom of a 96-well microtiter plate. A primary layer of alloantisera is first reacted against rat target cells. The secondary antibody, peroxidase conjugated antirat IgG, is then added to each well and serves as the second sandwich layer. Optimal reagent concentrations were determined by serial dilution analysis of various cell concentrations and secondary antibody dilutions. It was found that 200,000 cells per well was the optimal target cell concentration. However, 100,000 cells per well was also sufficient to run the assay with acceptable performance characteristics. Even lower cell concentrations of 10,000 and 20,000 cells/well, although not optimal, also produced acceptable results. Secondary antibody concentration with respect to the optimal cell concentration was determined to be 1:500. At 200,000 cells per well and a 1:500 secondary antibody dilution, the assay presented excellent coefficients of determination and high positive to negative ratios. The reaction was found to be very sensitive in yielding high antibody titers with low background levels and could be defined mathematically as a linear-log function. Titers of multiple unknown alloantibody samples were easily and accurately predicted in an automated manner by regression analysis form known standards. This immunoassay will be useful in studies of cell surface determinant expression and quantitation of antibodies reactive to such markers.

Postoperative abdominal wall defects with enterocutaneous fistulae.

BACKGROUND: Abdominal wall dehiscence with an associated enterocutaneous fistula is a surgical complication with high morbidity and mortality. Management of the abdominal wall defect is complicated by the continued emergence of liquid bowel contents. PATIENTS AND METHODS: Large abdominal wall wounds of 10 patients with postoperative abdominal wall dehiscence and active enterocutaneous fistulae were managed with early skin grafting directly onto the granulated abdominal viscera. RESULTS: Skin graft take averaged 93 +/- 12%, and there were no perioperative complications related to the skin grafting procedure. Overall mortality was 1 out of 10 patients. Enterocutaneous fistula output did not prove overly injurious to the skin grafts. Wound care was simplified in all but 1 patient with fitting of an ostomy appliance. CONCLUSION: Temporary abdominal wall wound closure with skin grafts improved patient comfort and simplified wound care in a staged reconstructive approach to this surgical complication.

Use of regression analysis and flow cytometry for determining levels of mixed semiallogeneic immune chimerism.

It has been shown that tolerance or specific immunologic nonresponsiveness in various lymphohemopoietic transplant models can be associated with the development of mixed lymphoid chimerism. As a specific example, composite tissue (limb) allografts were studied as a model for vascularized bone marrow transplantation (VBMT) and it was demonstrated that development of stable cellular immune chimerism is associated with long-term allograft survival. Recently, studies were initiated using a new parental to hybrid VBMT model, but the detection of donor cells is complicated, due to the fact that they share one parental allotypic determinant. Therefore, regression analysis with a flow cytometric immunofluorescent staining assay was evaluated for the assessment of cellular lymphoid chimerism in donor parental to hybrid (P-->F1) lymphohemopoietic transplant models. Standard curves consisting of known mixed populations of parental donor (Lewis, LEW) and hybrid host F1 (Lew x BN, LBN) lymphocytes were established. Standard curves were analyzed by linear regression statistics and excellent coefficients of determination (r > .881) were obtained for all standard curves. A highly statistically significant (p < .016) linear relationship between level of donor cell chimerism (independent variable) and percent stained (dependent variable) was determined. The technique was then evaluated using the parental to hybrid VBMT model. Levels of donor LEW lymphoid chimerism in all VBMT LBN recipients were successfully assessed by regression analysis and inverse prediction using distinct recipient allodeterminant markers. In conclusion, this technique was proven to be reliable and accurate for the detection of of chimerism in parental to F1 lymphohemopoietic allograft models.

Immune considerations in tissue engineering.

This article discusses the interplay between manmade living tissues and nature's given ones. Through a multidisciplinary approach, new strategies are outlined to control early inflammatory biochemical events, cell migration and development, and matrix deposition, all of them directed toward the acellular component of tissue-engineered constructs. The construct's resident nonautologous cells are targeted by cell- and humoral-mediated reactions, and modulatory strategies are applied to arrest rejection. The fate of ex vivo virally transfected, genetically engineered cells also is reviewed.

Emerging approaches to the tissue engineering of fat.

The eventual development of tissue-engineered fat equivalents for reconstructive and augmentation purposes will be most welcome by nearly every surgical discipline and prove to be especially useful for plastic surgeons. The clinical applications for which tissue-engineered fat will be particularly useful are vast and varied and can be loosely categorized into reconstructive, cosmetic, corrective, and orthotic indications. In this article, the authors discuss the emerging tissue-engineering strategies for fat, including the procurement of autologous cells, cell growth and differentiation, implantation and engraftment, polymer scaffolds, and implant integration and histogenesis.

Graft-versus-host disease in limb transplantation: digital image analysis of bone marrow and TGF-beta expression in situ using a novel 3-D microscope.

A subpopulation of parental to hybrid VBMT recipients developed characteristic clinical and histopathologic manifestations of GVHD. These changes are similar to those seen in human GVHD secondary to bone marrow transplantation. Human GVHD also manifests itself in an acute and chronic manner. Only a minority (30% to 40%) of animals developed lethal GVHD in our model. Those animals developing GVHD had a significantly (P < .0001) higher expression of TGF-beta in situ compared to the tolerant subpopulation. The differential expression of TGF-beta may represent an important mechanism of immune dysregulation associated with GVHD in CTA recipients.

Vascularized bone marrow transplantation (VBMT): induction of stable mixed T-cell chimerism and transplantation tolerance in unmodified recipients.

In this preliminary report, our model of VBMT across a semiallogeneic barrier consistently brings about antigen-specific host tolerance with absence of GVHD in the majority of recipients. No immunologic or radiologic intervention was utilized. These results emphasized a potentially important mechanism for low-level stable mixed lymphoid chimerism (SMLC) in tolerance induction, independent of immune suppressive effects due to irradiation or immunopharmacologic intervention.

Quantitative facial motion analysis after functional free muscle reanimation procedures.

The purpose of this study was to evaluate the success of functional free muscle transfer in patients with chronic facial paralysis using a recently developed quantitative method known as the maximum static response assay of facial motion. A retrospective review of a single surgeon series of six patients with longstanding facial paralysis was performed. The maximum static response assay was performed on all patients preoperatively and serially during the postoperative period. Twenty-seven patients (54 sides) with normal facial function were also evaluated and served as controls. The contralateral normal side in those patients with unilateral facial paralysis (n = 4) also served as a control. Movement of the modiolus during smile was recorded in the x axis and y axis. To determine net smile movement, the vector of movement was calculated by means of the Pythagorean theorem. Vectors were then defined mathematically by calculating direction and magnitude. The average direction of the vector during smile for the normal control population was 58.3 degrees (range 32.5 to 83.1 degrees) from the horizontal through the modioli, and the average magnitude was 10.6 mm (range 4.2 to 20.1 mm). The average preoperative direction for the reanimated sides was 176.8 degrees with a range of 83.3 to 225 degrees. Patients with bilateral paralysis (n = 2) were excluded for calculation of the vectors on the normal contralateral side. The average preoperative direction for the normal contralateral side in patients with facial paralysis was 58.3 degrees with a range of 48.2 to 68.4 degrees. Postoperatively, the average direction of the vector during smile for the reanimated sides improved to a value of 77.6 degrees with a range of 45.7 to 113.8 degrees. The average change in direction of the preoperative reanimated side compared with the postoperative reanimated side was significant (p = 0.01). Postoperatively, the average direction of the vector for the contralateral normal sides was 43 degrees with a range of 11 to 57.2 degrees. The change in direction for the contralateral normal side was not significant (p = 0.18). The average magnitude of the reanimated side improved from a non-anatomic 2.8 mm preoperatively (range 0.8 to 6.8 mm) to an anatomic 4.9 mm postoperatively (p = 0.02). The contralateral normal side magnitude decreased from 9.4 mm (range 7.3 to 11.6 mm) preoperatively to 5.7 mm (range 3.8 to 7.7 mm) postoperatively (p = 0.006). More specifically, the absolute change in movement on the reanimated side during smile for the x axis and y axis was 2.3 mm (p = 0.05) and 4.0 mm (p = 0.002), respectively. This corresponded to an absolute change in the magnitude of the vector of 4.6 mm in an anatomic direction. On the contralateral side the absolute change in magnitude during smile from preoperative to postoperative for the x axis and y axis decreased by 1.5 mm (p = 0.13) and 5.3 mm (p = 0.05), respectively. This reflected an absolute change in the magnitude of the vector of 5.5 mm. Functional free muscle transfer in patients with chronic facial paralysis resulted in anatomic recovery of motion in the majority of patients in this series. The maximum static response assay can be used to objectively assess the results of facial reanimation.

A novel device for the simple and efficient refinement of liposuctioned tissue.

In short, our device allows a surgeon who is harvesting adipose tissue for autologous fat transplantation to immediately, easily, efficiently, and sterilely isolate adipose tissue from the unwanted waste components that are associated with primary liposuction effluent. It does so by "trapping" the fat tissue contained within raw liposuction effluent. Once the tissue fraction has been separated, the device design then allows for direct implantation or subsequent washing/rinsing of the tissue with saline/buffer of choice in preparation for tissue reimplantation.

Composite tissue allografts in rats: IV. Graft-versus-host disease in recipients of vascularized bone marrow transplants.

This laboratory has used a composite tissue allograft model as a vehicle for studies on a new type of bone marrow transplant, the vascularized bone marrow transplant. The model consists of a rat hind limb transplant that incorporates integumentary musculoskeletal, and lymphopoietic tissues. These transplants, in comparison with conventional marrow transplants, have the advantage of providing a syngeneic microenvironment and immediate engraftment of both mature and progenitor hemopoietic cells at the time of transplantation. The characteristics of graft-versus-host disease were studied in this model. Lewis X Brown Norway F1 (LBN RT-1(1+n)) rats received hind limbs from Lewis (LEW RT-1(1)) donors (n = 19). Animals were observed daily for signs of graft-versus-host disease. Necropsies were performed. A minority of animals developed lethal disease (7 of 19 recipients) and demonstrated cachexia with concomitant histopathologic changes of the disease. Acute and chronic groups emerged with distinct clinical courses, which are similar to other models of this disease. Recipients of vascularized bone marrow transplants (limb transplants) showed clinical and histopathologic changes of the disease. The transplants may be used as a model of graft-versus-host disease in humans. Most interestingly, the transplant has a lower incidence of disease compared with other methods of bone marrow transplantation and represents an alternative to conventional bone marrow transplantation, which deserves further exploration. It may be possible to develop a new technique for bone marrow transplantation based on this surgical approach. It is proposed that the transfer of vascularized blocks of bone/marrow into prospective recipients as opposed to cellular bone marrow transplants may be preferable.

Computer-assisted anthropometry for outcome assessment of cleft lip.

Anthropometry and clinical examination best evaluate the morphology of repaired cleft lip and nose. An original, accurate, and practical image analysis of the lip and nose, which takes advantage of the mathematic, geometric, and organizational capabilities of public domain NIH-Image software (http://rsb.info.nih.gov/nih-image/), has been developed and tested over the past 6 years. A modified structured physical examination form that complements this analysis is under study. Accuracy of NIH-Image-based anthropometry was compared with direct measurements of 22 linear distances on the lip and nose. Twenty-five sets of direct measurements were taken, prospectively, on 15 children with repaired cleft lip over a 6-year period. The results were submitted to regression analysis. Then, relevant lip and nasal tip aesthetics were evaluated by the measuring capabilities of NIH-Image to create a quantitative assessment tool. For each episode, 15 possible faults were weighted, according to aesthetics and deformity, to provide an adverse score. The sum of the 5 lip scores, 10 nose scores, and combination gave respective grades. The analysis was modified to stratify congenital deformity to relate severity of disease to outcome. This analysis was applied to digitized images of 19 consecutive children, immediately prior to repair of complete unilateral cleft lip and nose, at the time of palate repair, and annually from the age of 3 to 6 years. There were 19 NIH-Image-based measurements of the congenital deformity and 35 measurements of surgical results; four children had three sets of records, eight had two sets, and seven had one set Descriptive statistics were applied. Following 556 paired direct and computer-assisted measurements, exceptional linear correlation was shown with a Pearson R coefficient of 0.96. The best correlation was lines within the plane of the camera lens, with the average difference ranging between 0.025 and 0.997 mm. Visual inspection of frontal and submental photographs of excellent, good, and poor results substantiates the ability of this analysis to quantify and grade a spectrum of relevant cleft lip and nasal anatomy. For these 19 patients, there was a broad range of performance scores, approximating a normal distribution. The mean of the NIH-Image-based analysis scores, 16.91, was a (very) good grade. A single standard deviation of 6.88 extended up into excellent and down to fair. The congenital analysis indicated a range of deformity. Comparing deformity with outcome, simple regression analysis had a coefficient of determination (R2) of 0.223, indicative of a weak positive relationship. An accurate and practical morphologic computer-assisted outcome assessment of repaired cleft lip and nasal deformity has been developed. There is a weak direct correlation between severity of deformity and outcome. Testing in multiple clinics is warranted.

Site-specific immunosuppression: mechanisms of cellular immunosuppression that are operative at local and systemic levels.

The cellular mechanisms by which topical cyclosporine A (tCsA) induces site-specific immunosuppression were investigated. Experiments were designed to elucidate how cyclosporine A (CsA) suppresses activated immunocytes in animals that are undergoing local alloactivation and concomitant tCsA immune suppression. Lewis rats received dual Lewis x Brown Norway rat skin allografts; the rats were treated with systemic CsA (sCsA) at 8 mg/kg/day for 10 days after grafting and then tCsA and vehicle thereafter. CsA added to mixed lymphocyte reactions 24 hours after culture initiation modeled the local effects of CsA on alloactivated immunocytes, and tCsA in conjunction with limited sCsA prolonged local skin allograft survival. CsA inhibited both antigen-specific and nonspecific activated alloresponses of immunocytes from animals that had received allografts and that underwent limited sCsA treatment only in a dose-dependent manner. When tCsA had been applied, immunocyte responses to a nonspecific antigen were extremely CsA-resistant as compared with those induced by antigen-specific suppression. However, this nonspecific alloresponse was fully suppressible with the use of elevated CsA doses (66 microg/mL); thus alloresponding immunocytes were significantly more sensitive to CsA if they were challenged with the donor antigen and preexposed to limited sCsA followed by tCsA in vivo.