The inverse relationship between microvessel counts and tumor volume in breast cancer.

Angiogenesis has emerged as an indicator of metastatic potential in invasive breast cancer. Exponential tumor growth and the appearance of metastasis are observed as new microvessels form. We postulated that the relevance of angiogenesis would be enhanced if analyzed as a function of tumor volume rather than greatest diameter alone and that microvessel counts would proportionately increase as does volume. Since tumors are three-dimensional solids, volume was calculated using the formula for an ellipsoid, V = pi/6 (a x b x c). Sixty-four tumors < or = 2.5 cm were studied and analyzed in 5 mm incremental ranges. Mean microvessel counts did not vary significantly among these tumor size groups. However, analysis of microvessel counts as a function of tumor volume decreased from 947.1/cm3 (0-0.5 cm) to 18.1/cm3 (2.1-2.5 cm), a greater than 50-fold difference. High microvessel density in small cancers supports the notion of metastasis as an early event, making these small tumors perhaps ideal targets for antiangiogenic agents.

Neutrophil adhesion to vascular prosthetic surfaces triggers nonapoptotic cell death.

OBJECTIVE: To test the hypothesis that neutrophil adhesion to expanded polytetrafluoroethylene (ePTFE) and Dacron triggers cell death. SUMMARY BACKGROUND DATA: Vascular prosthetic infections are intransigent clinical dilemmas associated with excessive rates of death and complications. Impaired neutrophil function has been implicated in the infection of implanted cardiovascular devices. ePTFE and Dacron are potent neutrophil stimuli able to elicit activation responses such as reactive oxygen species production independent of exogenous/soluble agonists. Reactive oxygen species that are released into the medium when neutrophils are challenged by soluble agonists are known to cause self-destruction. The authors therefore sought to examine whether neutrophil adhesion to prosthetic graft materials decreases neutrophil viability by means of reactive oxygen species production. METHODS: Neutrophils were adhered to surfaces for up to 6 hours. Cell viability was monitored with propidium iodide staining and lactate dehydrogenase release. RESULTS: Within 6 hours of adhesion to ePTFE and Dacron, respectively, 59% +/- 11% and 44% +/- 5% (n = 7) of the neutrophils were stained by propidium iodide. Indistinguishable results were obtained with plasma-coated ePTFE and Dacron. In contrast, less than 2% of the neutrophils adherent to fibrinogen-, immunoglobin-, or fetal bovine serum-coated polystyrene surfaces for 6 hours were positive for propidium iodide. The increase in membrane permeability to propidium iodide was accompanied by a two- to threefold increase in lactate dehydrogenase release. Pretreatment of neutrophils with N-acetyl-L-cysteine, cytochalasin D, or cyclosporin A significantly reduced the number of propidium iodide-positive ePTFE and Dacron adherent neutrophils. CONCLUSIONS: Neutrophil adhesion to ePTFE and Dacron triggers a rapid nonapoptotic cell death. The effect of ePTFE and Dacron on neutrophil viability appears to be caused by reactive oxygen species production. The premature death of graft-adherent neutrophils provides a novel explanation of the defect in neutrophil bacterial killing associated with vascular prosthetic grafts.

Subtle differences in quality of life after breast cancer surgery.

BACKGROUND: Lumpectomy with axillary dissection (LAD) has taken its place alongside mastectomy (M) as the treatment of choice for stage I and II breast cancer. Its appeal is based on lessening disfigurement and thus improving quality of life. METHODS: We used the SF-36 Health Survey modified with ten questions relevant to breast cancer surgery to evaluate whether quality of life with LAD was better than with mastectomy in women with stage I and II disease. The additional questions addressed satisfaction with intimate relationships and sexuality, and explored impact on the way women dress, use bathing suits, hug people, are comfortable with nudity, and rate their sexual drive and sexual responsiveness. RESULTS: LAD was not associated with statistically significant better quality-of-life scores on any SF-36 questions, except vitality (P = .02). No differences were noted in the areas of intimacy and sexual satisfaction. LAD patients reported significant differences in matters of dress, use of bathing suits, hugging, comfort with nudity, and sexual drive compared to patients undergoing mastectomy. CONCLUSIONS: The SF-36 health survey detected few differences in quality of life measures between patients with LAD and those with mastectomy. However, LAD impacts favorably on the way women dress, on comfort with nudity, and on sexual drive.

Immobilized IgG and fibrinogen differentially affect the cytoskeletal organization and bactericidal function of adherent neutrophils.

The infection of a vascular prosthesis is a dreaded clinical outcome. Since fibrinogen (FBGN) and immunoglobulin (IgG) coat the implanted biomaterial surface, it is with these immobilized proteins that the neutrophil (PMN) interacts. This study tests the hypothesis that PMN are impaired in their ability to kill bacteria when bound to immobilized IgG or FBGN. Isolated human PMN were bound to FBGN or IgG and then left untreated or exposed to phorbol myristate acetate (PMA; 10(-7) M). PMN adhered loosely, but did not spread, onto FBGN. In contrast, PMN spread fully onto IgG, exhibiting polarized pseudopodia. PMA treatment induced spreading of the FBGN bound cells. Suspended and adherent PMN were incubated 1 h with 14C-labeled Staphylococcus aureus; then, phagocytosis was assessed by radioactive uptake or bacterial kill was determined by plating recovered bacteria and colony counting. Data were analyzed by unpaired t test. We observed that both the phagocytic and the killing ability of FBGN-bound PMN were similar to that of suspended PMN. Conversely, IgG-bound PMN displayed a 62 +/- 6% (P < 0.01) decrease in phagocytosis and 33 +/- 7% (P < 0.05) reduction in kill vs suspended cells. PMA induced a 74 +/- 6% (P < 0.01) reduction in phagocytosis and 68 +/- 5% (P < 0.01) reduction in kill of bacteria for PMN bound to FBGN with no further effect on IgG-bound PMN. Using fluorescent vital dyes and confocal microscopy we determined that 33% fewer PMN were engaged in phagocytosis when bound to IgG vs FBGN. We conclude that Fc receptor ligation by immobilized IgG or PMA treatment of the FBGN-adherent PMN triggers cell spreading and reduced bactericidal activity. These results indicate that excessive cytoskeletal organization may impair the ability of PMN to kill bacteria and result in vascular graft infections.

The biologic role of B2 integrins in the host response to expanded polytetrafluoroethylene.

We hypothesized that the adherence of leukocytes to a vascular graft surface is mediated by B2 integrins. We studied integrin expression and monoclonal antibody inhibition of peripheral blood leukocyte (PBL) binding in vitro and CD11b expression in vivo. Human PBL were grown on ePTFE in culture and labeled with monoclonal antibodies to CD11a, -b, -c, and/or CD18. Percentage of monoclonal antibody binding and inhibition of leukocyte adherence was studied for up to 30 min using flow cytometry. ePTFE segments were implanted subcutaneously in SH-1 mice and PBL harvested 4 days later. PBL binding of monoclonal antibodies against CD11b was measured using flow cytometry. CD18 was constitutively expressed and CD11a decreased over time. CD11b increased from 41 to 62% and CD11c increased transiently (P < 0.003, P < 0.005). Inhibiton of PBL adherence was greatest by CD11b (34%) and CD11b + CD18 (57%) (P < 0.005, P < 0.0025). Implanted ePTFE caused a fourfold increase in PBL monoclonal antibody binding of CD11b (P < 0.001) compared to sham, Staphylococcus aureus alone, and combination of ePTFE and S. aureus. In conclusion, leukocyte integrins play a central role in the interaction between PBLs and ePTFE as measured by binding of monoclonal antibodies and inhibition of PBL adherence. This is also true in vivo since PBL increase CD11b expression four times when ePTFE is compared to sham. These observations indicate a potential role in vivo for B2 integrins in vascular prosthetic infections.

Three dimensional staging of breast cancer.

PURPOSE: Breast cancers are three dimensional solids but very few are spherical. We hypothesized that calculations based on the greatest diameter would not accurately reflect tumor volume and that three dimensional measurements would affect tumor staging. MATERIALS AND METHODS: 165 invasive carcinomas measuring 2.5 cm or less and having three measured diameters (a > or = b > or = c) noted were evaluated. Tumor volume was calculated using four geometric models: the spherical 4/3 pi (a/2)3, prolate spheroid 4/3 pi (a/2) (c/2)2, oblate spheroid 4/3 pi (a/2)2 (b/2), and ellipsoid 4/3 pi (a/2 x b/2 x c/2). The ellipsoid correctly determined the volume for any tumor shape. All cases were stratified according to the TNM staging system. Differences in mean volume calculated as a sphere and ellipsoid for each tumor subclass were analyzed using Student's T test. The reclassification of tumors by the ellipsoid formula was determined. RESULTS: Seventy-six (46.1%) had tumors with three different diameters while only six (3.6%) were true spheres having three identical diameters. Mean tumor volume analysis of T1a, T1b, T1c, and T2 tumors demonstrated a statistically significant overestimation of volume when utilizing the sphere formula instead of the ellipsoid formula (p < 0.05). The differences in volume were more dramatic as the diameters increased. A total of 41 tumors were moved into smaller T subclasses including 10 node positive patients. CONCLUSIONS: Tumor volume analysis demonstrates that use of only the greatest diameter poorly reflects the true volume of a lesion and consistently overestimates volume. The ellipsoid formula accurately calculates volume for these three dimensional tumors and when utilized has significant relevance to staging small invasive breast cancers.

Endothelial cells adherent to expanded polytetrafluoroethylene express the intercellular adhesion molecule-1.

BACKGROUND: The foreign body reaction caused by implanted biomaterials is not fully characterized. To evaluate the effect of an expanded polytetrafluoroethylene (ePTFE) surface on the binding of monoclonal antibodies against the intercellular adhesion molecule (ICAM-1) by adherent endothelial cells, an in vitro bioassay was developed. STUDY DESIGN: Human saphenous vein endothelial cells (HSVEC) and human umbilical vein endothelial cells (HUVEC) were grown to confluency on fibronectin-pretreated ePTFE and on fibronectin-pretreated polystyrene culture flasks used as controls. Cells were assayed for size, density, viability, and binding of anti-ICAM-1 antibodies. Human peripheral blood leukocytes (PBLs) were then cocultured with HUVEC adherent to ePTFE and control polystyrene and HUVEC were assayed for the binding of anti-ICAM-1 antibodies. RESULTS: The adherence of HSVEC and HUVEC to ePTFE resulted in a twofold increase in the binding of monoclonal antibodies against ICAM-1 compared with controls at 48 hours (p < 0.005). The addition of PBLs to the adherent HUVEC resulted in a four- to eightfold increase in anti-ICAM-1 antibody binding on ePTFE and polystyrene, respectively, at 24 hours. CONCLUSIONS: Adherence of endothelial cells to ePTFE results in an increase in the binding of anti-ICAM-1 antibodies. Coculture of adherent HUVEC and PBLs results in a marked increase in the number of cells binding ICAM-1 antibodies. These data support the conclusion that ePTFE is associated with the activation of adherent cells, which is one aspect of a multicellular inflammatory response.

Fc gamma RII, Fc gamma RIII, and CD18 receptors mediate in part neutrophil activation on a plasma coated expanded polytetrafluoroethylene surface.

BACKGROUND: A biomaterial-induced polymorphonuclear neutrophil (PMN) defect may predispose the implanted vascular graft to infection. PMNs bind, activate, and undergo morphologic changes when exposed to uncoated or plasma coated expanded polytetrafluoroethylene (ePTFE) surfaces. The purpose of this study was to investigate whether the CD18 integrin receptor or the immunoglobulin receptors Fc gamma RII and Fc gamma RIII mediate either PMN binding or activation on ePTFE. METHODS: PMN binding and activation were determined after incubation of these cells on human immunoglobulin (IgG) or fibrinogen coated surfaces and uncoated or plasma coated ePTFE. PMN activation was measured by using the ferricytochrome reduction assay. Binding was determined with chromium 51-labeled PMNs. To block the Fc gamma RII, Fc gamma RIII, and CD18 receptors, PMNs were preincubated with the monoclonal antibodies (mAbs) IV.3, 3G8, and IB4, respectively. Irrelevant isotype matched mAbs were used as control. RESULTS: Monoclonal antibody IB4 inhibited binding of activated PMNs to fibrinogen coated surfaces. Binding to IgG was affected by either mAb IB4 or IV.3, but the greatest degree of inhibition was achieved when mAbs IB4 and IV.3 were used in combination. IgG-induced activation was partially inhibited by mAb IV.3 but was fully inhibited by a combination of mAbs IB4 and IV.3 The mAbs did not affect PMN binding to uncoated or plasma coated ePTFE, nor was PMN activation on the uncoated ePTFE surface inhibited. PMN activation on the plasma coated ePTFE surface was, however, partially inhibited by the combination of mAb IB4 with either mAb IV.3 or 3G8. CONCLUSIONS: A synergistic interaction between the PMN Fc gamma RII receptor and the CD18 integrin receptor accounts for surface bound IgG-induced cell activation. Both receptors also play a role in mediating PMN activation on the plasma-coated ePTFE surface.

The future of general surgery characterized by general surgical chief residents.

The future of any specialty in medicine depends in large part on the career choices of its trainees. We obtained data from General Surgery Chief Residents who finished their training in 1990 and compared their attitudes to a similar survey conducted by us in 1985. According to those responses, residents in general surgery continue to choose fellowship training at a time when they no longer perceive an oversupply of general surgeons. This has broad implications for the future of general surgery as a specialty that requires significant change in the structure of our residency programs.

Patency of a small vessel prosthesis bonded to tissue plasminogen activator and iloprost.

To test the thrombosis resistance of a vascular prosthesis coated with antithrombogenic agents, we evaluated a small vessel prosthesis of expanded polytetrafluoroethylene (ePTFE) implanted in the rat aorta and removed 1 week following surgery. Control grafts consisted of 1 mm internal diameter ePTFE. Experimental grafts consisted of 1 mm internal diameter ePTFE noncovalently bonded to tissue plasminogen activator (tPA) and iloprost using the surfactant tridodecylmethylammonium chloride. After 1 week the grafts were harvested, patency was determined, and histologic specimens were prepared for electron microscopy. Six of 10 control grafts were thrombosed, whereas 9 of 10 tPA-iloprost-bonded grafts were patent (p < 0.03). Of concern, there was an unexpectedly high mortality rate in the tPA-iloprost group compared to the control group among animals that died before completion of the study. Evaluation of the safety of these drugs must, therefore, be an early component of future experiments. Nevertheless, these studies indicate that a small vessel prosthesis bonded to tPA and iloprost may ameliorate some of the complications associated with early graft failure.

Role of adhesion molecules in leukocyte binding to endothelial cells adherent to vascular grafts.

BACKGROUND: The localization of leukocytes to vascular grafts is an essential part of healing and infection resistance. The mechanisms involved in this process are only partly understood. STUDY DESIGN: Human saphenous vein endothelial cells (HSVEC) were grown on control polystyrene culture ware and expanded polytetrafluoroethylene (ePTFE). The binding of monoclonal antibodies against the intercellular adhesion molecule (ICAM-1) and the E-selectin by adherent HSVEC was determined by flow cytometry. Peripheral blood leukocytes (PBL) were cocultured with HSVEC adherent to ePTFE and leukocyte binding was determined with and without the addition of a protein kinase C inhibitor. RESULTS: HSVEC adherent to ePTFE constitutively bound anti-ICAM-1 antibodies, which were attenuated by the protein kinase C inhibitor, H-7. HSVEC adherent to ePTFE bound significantly greater numbers of leukocytes than those on control (58 versus 41 percent, p < 0.05). Incubation with H-7 decreased leukocyte binding to HSVEC significantly (p < 0.005). Coculture of PBL with HSVEC adherent to ePTFE caused a tenfold increase in binding of anti-E-selectin antibodies (p < 0.0005). CONCLUSIONS: These data indicate that PBL binding to HSVEC adherent to ePTFE is, at least in part, ICAM-1 to HSVEC adherent to ePTFE is, at least in part, ICAM-1 and E-selection dependent.

Neutrophil activation by expanded polytetrafluoroethylene is dependent on the induction of protein phosphorylation.

BACKGROUND: Polymorphonuclear leukocyte (PMN) activation after interaction with implantable surfaces has been previously reported. The purpose of this study was to examine the mechanism of PMN activation in response to expanded polytetrafluoroethylene (ePTFE). METHODS: To demonstrate PMN activation, the cumulative production of superoxide was measured on uncoated, plasma coated, or albumin coated ePTFE discs. Chromium 51-labeled PMNs were used to measure binding. Cell structure was examined by scanning electron microscopy. RESULTS: By 4 hours, PMN activation on either uncoated or plasma coated ePTFE was approximately 30% of phorbol 12-myristate 13-acetate-induced activation. Albumin inhibited PMN activation by ePTFE. No apparent correlation existed between chromium 51-labeled PMN binding and cell activation on the surfaces. Pretreatment of the cells with the protein kinase inhibitors bisindolylmaleimide or genistein resulted in marked inhibition of superoxide production on the uncoated and plasma coated ePTFE surfaces, whereas binding to these surfaces was not affected. PMNs spread on the uncoated surface and transmigrated into the plasma coated ePTFE surface. These effects of ePTFE on cell structure were inhibited by bisindolylmaleimide and genistein. CONCLUSIONS: ePTFE induced PMN activation, as measured by superoxide production, and changes in cell behavior are dependent on the activation of signaling pathways that involve protein phosphorylation events.

Multiple platelet surface receptors mediate platelet adhesion to surfaces coated with plasma proteins.

The interactions between platelets and plasma proteins previously shown to adhere to biomaterials were evaluated, using monoclonal antibodies (mAbs) against specific platelet surface glycoprotein (GP) receptors. Purified 51Cr-labeled human platelets in plasma-free medium were incubated with each of the following antibodies: mAb 10E5 [anti-GP IIb/IIIa; fibrinogen, von Willebrand factor (vWF), and fibronectin receptor]; mAb 6D1 (anti-GP Ib-IX; vWF receptor); mAb IV.3 (anti-Fc gamma RII; IgG receptor); polyclonal antiserum A108 or mAb BIIG4 (anti-GP Ic-IIa; fibronectin receptor). Antibody-treated platelets were added to microtiter wells coated with fibronectin, fibrinogen, vWF, IgG, vitronectin, albumin, or platelet-poor plasma (PPP). 51Cr-labeled platelet adhesion to matrix proteins was expressed as a percentage of that measured on PPP-coated surface. Platelets adhered to fibrinogen, fibronectin, vWF, or IgG immobilized on polystyrene. Limited binding to either vitronectin or albumin was detected. Binding to fibrinogen and IgG was blocked by mAb 10E5. Binding to IgG was also blocked by mAb IV.3. Binding to fibronectin, reduced in the presence of mAb 10E5, mAb BIIG4, or the polyclonal antiserum A108 alone, was further reduced by combined 10E5 and BIIG4 or 10E5 and A108. Neither mAb 10E5 nor 6D1 alone blocked adhesion to vWF; however, the combination of 10E5 and 6D1 significantly reduced platelet adhesion to this matrix. Finally, platelet adhesion to the plasma-coated surface was reduced by mAbs 10E5 and BIIG4. These results indicate that multiple adhesion receptors can mediate platelet adhesion to matrix proteins immobilized on surfaces.

Increases in type III collagen gene expression and protein synthesis in patients with inguinal hernias.

OBJECTIVE: The aim of this study was to determine if alterations in fibrillar collagen synthesis were associated with the development of inguinal hernias. SUMMARY BACKGROUND DATA: Previous work has suggested that alterations in connective tissue accumulation may play a functional role in the development of inguinal hernias. In particular, several investigators have suggested that alterations in collagen synthesis, causally related to connective disorders such as osteogenesis imperfecta, may also be responsible for the inguinal herniation that is markedly increased in such patients. This study was undertaken therefore to study collagen synthesis in patients with inguinal hernia in the absence of any other connective tissue disease. METHODS: Skin fibroblasts from 9 patients with hernias and 15 control individuals were radiolabeled with 3H-proline. Trypsin-chymotrypsin-resistant type I and III collagens were isolated and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis, and recovery was quantified by laser densitometry. Steady-state levels of alpha 1(I) and alpha 1(III) procollagen mRNAs were also determined by northern and slot-blot hybridization analysis. RESULTS: The alpha 1(I)/alpha 1(III) collagen ratios were shown to be 6.3 +/- 0.34 in fibroblasts from control individuals and 3.0 +/- 0.25 in fibroblasts from patients with inguinal hernias. This statistically significant difference (p < 0.0001) was caused by an increase in the secretion of alpha 1(III) procollagen from the fibroblasts of patients with hernias. A concomitant increase in the steady-state levels of alpha 1(III) procollagen mRNA was observed in total RNA isolated from the patients' fibroblasts. CONCLUSIONS: A constitutive and systemic increase in type III collagen synthesis may result in reduced collagen fibril assembly in the abdominal wall, eventually leading to the development of herniation. Although it is not yet clear what genetic factors are responsible for the elevation in type III collagen synthesis in patients with hernias, this study represents the first attempt to define individuals with an abnormality in collagen production that may be specifically related to herniation. A clearer understanding of the possible genetic factors that influence the pathophysiology of this disease will be important to improve the treatment of patients in whom inguinal hernias develop.

Integrins, adhesion molecules and surgical research.

Integrins and adhesion molecules are a group of closely related glycoproteins expressed by a wide variety of cells. In vitro experiments have shown these families of cell surface receptors to be essential to the interactions between the cytoskeleton and the extracellular matrix. Although the designations of families of integrins and adhesion molecules is complex, this review attempts to present an organization in which the result is to provide surgeons with an understanding of their structure and function. This is particularly important because integrins and adhesion molecules have a critical role in such diverse processes as embryogenesis, inflammation, wound healing, thrombosis, transplantation and oncogenic transformation. It is, therefore, highly probable that they will be critical to future advances in surgical research and ultimately in surgical practice. It should be remembered that this area of investigation is so dynamic that the interval involved in publication of this article will in itself be characterized by a plethora of publications in the basic science and surgical literature, which warrants careful attention.

Mammographic changes following biopsy and lumpectomy-breast irradiation.

Mammographic architectural distortion occurred at the operative site in 86 percent of lumpectomy patients and in 34 percent of biopsy patients during the first year (P < 0.001). These changes can mimic carcinoma and may be slow to resolve.