An enzyme-mediated assay to quantify inoculation volume delivered by suture needlestick injury: two gloves are better than one.

BACKGROUND: Acquiring a blood-borne disease is a risk of performing operations. Most data about seroconversion are based on hollow-bore needlesticks. Some studies have examined the inoculation volumes of pure blood delivered by suture needles. There is a lack of data about the effect of double-gloving on contaminant transmission in less viscous fluids that are not prone to coagulation. STUDY DESIGN: We used enzymatic colorimetry to quantify the volume of inoculation delivered by a suture needle that was coated with an aqueous contaminant. Substrate color change was measured using a microplate reader. Both cutting and tapered suture needles were tested against five different glove types and differing numbers of glove layers (from zero to three). RESULTS: One glove layer removed 97% of contaminant from tapered needles and 65% from cutting needles, compared with the no-glove control data. Additional glove layers did not significantly improve contaminant removal from tapered needles (p > 0.05). For the cutting needle, 2 glove layers removed 91% of contaminant, which was significantly better than a single glove (p = 0.002). Three glove layers did not afford statistically significant additional protection (p = 0.122). There were no statistically significant differences between glove types (p = 0.41). CONCLUSIONS: With an aqueous needle contaminant, a single glove layer removes contaminant from tapered needles as effectively as multiple glove layers. For cutting needles, double-glove layering offers superior protection. There is no advantage to triple-glove layering. A surgeon should double-glove for maximum safety. Additionally, a surgeon should take advantage of other risk-reduction strategies, such as sharps safety, risk management, and use of sharpless instrumentation when possible.

Review of vascularized bone marrow transplantation: current status and future clinical applications.

In this review, we examine the applicability of the vascularized bone marrow transplant (VBMT) as an alternative to conventional bone marrow transplantation (BMT). As a new surgical approach, the VBMT is unique by transplantation of the stromal environment that eliminates the need for an engraftment period, provides critical signaling and modulatory functions, and may potentiate tolerance induction. Thus far, VBMT studies have demonstrated an absence of graft-versus-host disease (GVHD) and robust engraftment into nonmanipulated as well as irradiated recipients with evidence of immunological tolerance. Further investigation is needed to determine the applicability of VBMT as an alternative to BMT.

Pulse doppler and M-mode to assess viability of cardiac allografts using heterotopic femoral heart transplantation in rats.

Noninvasive assessment of heterotopic heart transplants using Doppler echocardiography was first described in two patients by Allen at Stanford in 1981. Since then, numerous experiments studying heterotopic heart transplantation in humans and large animals have confirmed its utility by employing either an intra-abdominal or cervical model. In rats, however, prior research investigating intra-abdominal heterotopic hearts has showed echocardiography to be ineffective. We have recently developed a new technique for heterotopic femoral heart transplantation in rats, which employs the novel use of trans-femoral echocardiography. Therefore, our goal was to re-examine the efficacy of echocardiography for detection of graft rejection.

A new modified technique for heterotopic femoral heart transplantation in rats.

BACKGROUND: Abbott developed the first experimental accessory heart transplant rat model in 1964. This intra-abdominal model required a labor-intensive aortic anastomosis. In 1971, Heron modified the operation by using sutureless cervical vessel anastomoses. Rao and Lisitza developed a femoral heart accessory transplant model in 1985. Our goal was to improve this femoral model for the study of cardiac transplantation between both syngeneic and allogeneic rats. METHODS: ACI and Lewis rats weighing 150 to 350 g were used as donors and recipients (n = 12). The left common carotid and left pulmonary arteries were anastomosed to the femoral artery and vein in an end-to-end fashion, respectively. Improved modifications included the use of hemostatic vessel clips, heparinization of both donor and recipient, a ventricular prolene stay-suture for secure graft placement, and transfemoral echocardiography (TFE). Total operative time averaged 61 +/- 12 minutes. RESULTS: Femoral accessory transplanted hearts (FATHs) allowed easier pulse palpation and access for TFE versus previously described cervical and intra-abdominal models. This modification allows precise detection of acute graft rejection (AGR) and is defined as absent ventricular contraction in the presence of anastomostic patency. CONCLUSIONS: Our new modified technique for heterotopic femoral heart transplantation in rats is a relatively easily learned and reproduced procedure that allows superior allograft access for palpation and improved echocardiographic assessment. Femoral heterotopic heart transplantation remains an effective model for allograft transplantation study.

Absence of graft-versus-host disease in the isolated vascularized bone marrow transplant.

An isolated vascularized bone marrow transplant (iVBMT) model was developed to study the contribution of the bone marrow component in a composite tissue allograft. We hypothesized that the iVBMT would be functional and cause graft-versus-host disease (GVHD) in a fraction of the recipients. Lewis iVBMT grafts were transplanted to Lewis-Brown Norway recipients. Animals were sacrificed at various times from 1 to 14 weeks. Polymerase chain reaction for microchimerism was performed on the host's marrow. No animals exhibited signs of GVHD at death. Histologic examination of the grafts showed a normal mix of hematopoietic and fatty elements and appeared to be functional. Tissues usually affected-tongue, ear, liver, and gut-also showed no evidence of disease. Polymerase chain reaction demonstrated microchimerism in both groups. These findings suggest that the vascularized bone marrow within a composite tissue allograft is not the component that causes GVHD; rather, it may serve an immunomodulatory function for tolerance induction.

An extraperitoneal isolated vascularized bone marrow transplant model in the rat.

An isolated vascularized bone marrow transplant (iVBMT) model was previously developed in the rat to specifically study the role of bone marrow and its environment in a composite tissue allotransplant. An extraperitoneal model was successfully created to avoid laparotomy and cross-clamping of the great vessels. The extraperitoneal iVBMT model consisted of a left donor femur that was harvested with its nutrient vessels, anastomosed to the right femoral vessels in a syngeneic host, and then placed subcutaneously in the abdominal wall. At explant, the graft vessels were grossly patent, and histology of the graft bones showed a viable marrow compartment. Polymerase chain reaction demonstrated peripheral chimerism in the recipients. This model is technically simple with minimal morbidity in the recipient animals. By using the iVBMT, future studies across semiallogeneic and allogeneic barriers will help define the role of the bone marrow compartment in composite tissue allotransplants to potentially induce immune tolerance.

Engineering a composite neotrachea with surgical adhesives.

BACKGROUND/PURPOSE: Reconstructive surgery often is limited by the availability of normal tissue. Tissue engineering provides promise in the development of "artificial tissues." The purpose of this study was to test the efficacy and viability of the use of a biologic surgical adhesive TISSEEL in combining engineered bronchial epithelium with engineered cartilage. METHODS: Using isolated human cells, bronchial epithelium and mature cartilage were engineered. Using a contact adhesive technique, TISSEEL was used to biologically fuse the bronchial epithelium and the cartilage. The fused composite then was supported for 5 days in tissue culture. The mechanical properties of the adhesion were tested, and the construct was studied morphologically to assess viability of the cartilage and the bronchial epithelium. The bronchial epithelium showed a normal cell size (337.2 microm2) and epithelial thickness (46.47 microm). RESULTS: TISSEEL was effective in fusing the epithelium to the cartilage. The construct remained viable for 5 days in culture. There was no difference in the dimensions of the bronchial epithelium or the epithelial cells. Mechanical adhesion was achieved. CONCLUSIONS: Biologically compatible fibrin glue is an effective surgical adhesive that allows the tissue types to be fused while remaining viable and morphologically accurate. Surgical adhesives may show promise in the development of composite tissue development in the field of bioengineering.

VEGF upregulates Bcl-2 expression and is associated with decreased apoptosis in neuroblastoma cells.

BACKGROUND/PURPOSE: Both the expression of Bcl-2 and the amount of vascular endothelial growth factor (VEGF) are increased in neuroblastoma cells cocultured with hepatocytes. The authors hypothesize that VEGF upregulates Bcl-2 expression by the neuroblastoma cells and protects them from apoptotic stimuli. METHODS: To determine whether VEGF will induce Bcl-2 expression in neuroblastoma cells, the cells are plated with standard media (control) or media supplemented with VEGF. After 24 hours, Bcl-2 expression is measured. To determine whether VEGF protects neuroblastoma cells from apoptosis, the cells are subjected to tumor necrosis factor alpha (TNF-alpha) or serum starvation to induce apoptosis either with or without VEGF added to the culture media. The cells are collected and apoptosis measured using the deoxynucleotidyltransferase-mediated dUTP neck end labeling (TUNEL) method. RESULTS: VEGF increases Bcl-2 expression by 33% over cells cultured in standard media. Serum starving the tumor cells or adding TNF-alpha significantly increases the percentage of apoptotic cells. The addition of VEGF significantly protects the neuroblastoma cells from the apoptotic effects of both serum starvation and TNF-alpha. CONCLUSIONS: VEGF increases the expression of Bcl-2 and also abrogates TNF-alpha and serum starvation-induced apoptosis in neuroblastoma cells in vitro. VEGF may promote neuroblastoma survival not only through angiogenesis, but also by altering apoptosis and its regulating proteins.

Insulin-like growth factor-I protects neuroblastoma against starvation-induced apoptosis and is associated with increased Bcl-2 expression.

BACKGROUND/PURPOSE: Aggressive neuroblastomas avoid apoptosis and have increased expression of the antiapoptotic protein, Bcl-2. Insulin-like growth factor-I (IGF-I) is mitogenic and may promote tumor survival by inhibiting apoptosis. The authors hypothesize that IGF-I may protect neuroblastoma cells from apoptosis by upregulating their Bcl-2 expression. METHODS: Human neuroblastoma cells (IMR-32) are cultured, and 3 experimental groups are established: 1 group with cells cultured in standard growth media (control), 1 with cells grown in serum-depleted media (starvation), and 1 with neuroblastoma cells cultured in starvation media plus IGF-I. The cells are harvested at 14 and 24 hours, and cytospin slides are made. Bcl-2 expression is measured by immunohistochemistry. Apoptosis is detected with the TUNEL method. RESULTS: Bcl-2 expression is decreased 90% in the serum starved neuroblastoma cells. In addition, apoptosis is 150 times higher in the starved neuroblastoma cells. These changes are abrogated by the addition of IGF-I, where apoptosis is decreased 50% and Bcl-2 is 14-fold higher in the IGF-I-treated group. These changes are most apparent at 24 hours. CONCLUSIONS: IGF-I protects neuroblastoma cells from apoptosis and increases Bcl-2 expression. Growth factors may have a direct role in promoting tumorigenesis by inducing the expression of antiapoptotic proteins by the tumor.