Carolyn M. DeRosa

Angiogenesis is enhanced in ischemic canine myocardium by transmyocardial laser revascularization

OBJECTIVES This study sought to test whether transmyocardial laser revascularization (TMLR) stimulates angiogenesis in an animal model of chronic ischemia. BACKGROUND TMLR relieves angina and may also improve blood flow in patients who are not candidates for traditional therapies. The mechanisms of these benefits are not fully defined. METHODS Ischemia was created in 14 dogs by proximal left anterior descending coronary ameroid constrictors. TMLR was performed in the anterior wall (approximately 1 channel/cm2) of seven dogs; the remaining dogs served as the ischemic control group. Myocardial blood flow was measured (colored microspheres) at rest and during chemical stress (adenosine) in the acute setting and after 2 months. RESULTS TMLR did not influence blood flow in the acute setting. After 2 months, resting blood flow increased comparably in the anterior wall in both groups to approximately 80% of normal. However, the TMLR-treated dogs demonstrated an approximately 40% increase in blood flow capacity during stress in the ischemic territory compared with untreated dogs (left anterior descending coronary artery/left circumflex coronary artery flow 0.53+/-0.16 in the control group vs. 0.73+/-0.08 in TMLR animals, p < 0.05). Vascular proliferation, assessed by bromodeoxyuridine incorporation and proliferating cell nuclear antigen positivity in endothelial and smooth muscle cells was about four times greater in the TMLR group than in the control group (p < 0.001). The density of vessels with at least one smooth muscle cell layer was approximately 1.4 times greater in the myocardium surrounding the TMLR channel remnants than in control ischemic tissue (p < 0.001). CONCLUSIONS In this canine model of chronic ischemia, TMLR significantly enhances angiogenesis as evidenced by the increased number of vessels lined with smooth muscle cells, markedly increased vascular proliferation and increased blood flow capacity during stress.

Use of aprotinin in LVAD recipients reduces blood loss, blood use, and perioperative mortality

Aprotinin, a bovine protease inhibitor, has been used extensively in patients undergoing cardiac surgical procedures in an effort to minimize blood loss and prevent the complications associated with blood replacement. We sought to evaluate the effect of aprotinin on postoperative blood loss, renal function, and the incidence of right ventricular failure in patients undergoing placement of a TCI Heartmate left ventricular assist device as a bridge to cardiac transplantation. Retrospective data analysis in 142 patients (42 receiving aprotinin and 100 untreated) demonstrated that the use of aprotinin was associated with a significant decrease in postoperative blood loss (p = 0.019) and in the intraoperative packed red blood cell transfusion (p = 0.019) and total blood product (p = 0.016) requirements. A transient, yet significant, increase in the postoperative creatinine level in the aprotinin group (p = 0.0006), but not in blood urea nitrogen level (p = 0.22), was noted. Interestingly, we noted an association between blood loss and the subsequent development of right ventricular failure; patients who required a right ventricular assist device bled significantly more than did those who did not suffer right ventricular failure (p = 0.02). Additionally, aprotinin recipients benefited by a reduction of nearly one half in the incidence of the need for a right ventricular assist device. The incidence of perioperative mortality was reduced in those receiving aprotinin compared with that in untreated patients, (p = 0.05). We conclude that aprotinin is safe and effective in decreasing postoperative blood loss and intraoperative blood product requirements, and in reducing perioperative mortality in patients undergoing left ventricular assist device placement as a bridge to cardiac transplantation.

Physiology, histology, and 2-week morphology of acute transmyocardial channels made with a CO2 laser

BACKGROUND Transmyocardial revascularization with a CO2 laser appears to improve symptoms in patients with refractory angina. However, it remains controversial as to whether blood flow through the channels is the mechanism of benefit, especially in the acute setting. METHODS AND RESULTS Three protocols were used to test whether blood flows through transmyocardial CO2 laser revascularization channels. First, channels were made in excised, cross-perfused dog hearts (n = 5) using a CO2 laser (The Heart Laser; PLC Systems Inc, Milford, MA; 40 J/pulse) followed by ligation of the proximal left anterior descending coronary artery. Colored microspheres injected into the left ventricular chamber failed to detect any significant transmyocardial blood flow. In the second protocol (n = 4), laser channels were created in the left anterior descending artery territory, the left anterior descending artery was ligated, and the hearts were excised after 24 hours. Triphenyltetrazolium chloride staining revealed that no viable myocardium was detected around the laser channels in the ischemic myocardium. Finally, channels examined 2 weeks after creation in normal (n = 6) or ischemic (n = 4) myocardium did not maintain their original caliber but were invaded by granulation tissue, which included a large amount of smaller vascular spaces and vessels of various sizes. CONCLUSIONS Transmyocardial laser revascularization channels made with this CO2 laser did not provide acute myocardial perfusion or preserve myocardial viability in the face of acute ischemia. Channel morphology changes dramatically within the first 2 weeks. To the degree that these findings pertain to human myocardium, the results suggest that transmyocardial blood flow may not be the mechanism of benefit of this procedure, particularly in the acute setting.

Safety and efficacy of aprotinin under conditions of deep hypothermia and circulatory arrest

Aprotinin has been successfully used to reduce blood loss and blood product requirements in patients undergoing primary and reoperative cardiac operations. Its safety and efficacy during profound hypothermia and circulatory arrest have been questioned, however. A retrospective review compared 24 patients who received aprotinin during complex aortic procedures under profound hypothermia and circulatory arrest with 24 age-matched patients undergoing similar procedures without aprotinin. Activated clotting time was maintained at longer than 500 seconds (kaolin activating agent) or longer than 750 seconds (celite). We observed no statistically significant difference in the incidence of neurologic events (p not significant) or myocardial infarctions (p not significant), and there was a trend toward reduced in-hospital mortality rate in aprotinin-treated patients. A higher incidence of postoperative renal dysfunction was encountered in aprotinin-treated patients. Aprotinin recipients had a significant reduction in requirements for postoperative homologous erythrocytes (p = 0.01). We conclude that aprotinin may be safely and effectively used in patients undergoing deep hypothermia and circulatory arrest.

An immunohistochemical evaluation of progesterone receptor in frozen sections, paraffin sections, and cytologic imprints of breast carcinomas

Two monoclonal antibodies to progesterone receptor (PR), JZB39 and KD68, were used for the immunocytochemical visualization of PR in different kinds of breast cancer specimens including (1) cryostat sections of tumors frozen at −80°C; (2) paraffin sections of tumors fixed in formalin or in Bouins fixative for varying periods of time at room temperature or at 4°C; and (3) imprints and cryostat sections prepared from the tissue used for frozen section diagnosis and stored at −80°C after fixation in Zambonis solution. Sections of conventionally frozen specimens as well as imprints and cryostat sections stored for varying periods of time were stained with the peroxidase–antiperoxidase technique, whereas the avidin–biotin technique was used for paraffin sections. In all types of specimens the PR immunostaining was localized to the nuclei of carcinoma cells and displayed considerable heterogeneity both in intensity and in distribution of positive cells. Close correspondence was found between the different immunohistochemical techniques as well as between immunostaining and steroid‐binding assays. PR staining was more frequently positive in well‐differentiated than in moderately or poorly differentiated carcinomas, whereas no meaningful correlation was found between PR staining and extent of the disease. Similar results were obtained with the immunostaining of estrogen receptor in the same material using monoclonal antibodies H222 and D75P3γ. Thus, by choosing the technique that best suits the type of specimen available, it is possible to obtain valid information on the receptor status of any breast carcinoma, regardless of its size and clinical presentation.

Radio frequency transmyocardial revascularization enhances angiogenesis and causes myocardial denervation in canine model.

Transmyocardial revascularization (TMR) relieves angina and improves exercise tolerance in patients. Angiogenesis and myocardial denervation have been proposed as factors contributing to these benefits. To test whether radio frequency transmyocardial revascularization (RF‐TMR) enhances angiogenesis and causes myocardial denervation.

Antiproliferative effects of natural interferon beta alone and in combination with natural interferon gamma on human breast carcinomas in nude mice

SummaryNude mice bearing bilateral xenografts of human breast carcinoma cells (MCF-7 and BT20) were treated with 2 or 4 5-day cycles of intralesional (i.l.) injections of human natural interferon beta (nIFN-β) alone or in combination with human natural interferon gamma (nIFN-γ). The injections were administered to only 1 of the 2 tumors in each animal, thus making it possible to assess at the same time local therapeutic effects in the injected tumors and systemic effects in the contralateral ones. When n-IFN-β was used as a single agent only mild local antitumor effects and virtually no systemic effects were observed. In contrast, the combined administration of nIFN-β/nIFN-γ produced marked antiproliferative effects, presumably as a result of the synergistic action of type I and type II IFNs. These effects ranged from complete regression documented histologically in 2 MCF-7 tumors to varying degrees of growth inhibition with persistence of residual microscopic or grossly detectable tumor. Local effects were more pronounced than systemic effects. The therapeutic efficacy of nIFN-β proved to be greater than that of recombinant interferon beta (rIFN-β). In MCF-7 tumors nIFN-β appeared to be less effective than nIFN-α, whereas the opposite was true for BT 20 tumors.

Immunohistochemical assessment of estrogen and progesterone receptors in stored imprints and cryostat sections of breast carcinomas

A peroxidase-antiperoxidase technique was used to visualize estrogen and progesterone receptors in stored imprints and cryostat sections of breast carcinomas that were prepared at the time of biopsy or frozen section diagnosis. This was done to provide an alternate technique for the assessment of the receptor status of tumors that could not be adequately assayed with other biochemical or immunocytological methods. Fixation in Zambonis fixative followed by passage through cold methanol and acetone before storage at -80 C insured good preservation of the receptor proteins over extended periods of time (up to 56 weeks). Immunostaining of these stored preparations with monoclonal antibodies against estrogen receptor (H222) and progesterone receptor (JZB39 and KD68) showed a high degree of correspondence with immunocytochemical assays (ER-ICA and PR-ICA) and biochemical analysis. This technique is easy to perform and provides reliable information, even in tumors that are too small and/or ill defined to permit separate sampling for receptor assays.

Applicability of Monoclonal Antibodies Against Estrogen and Progesterone Receptors in the Immunocytochemical Evaluation of Breast Carcinomas

The development of monoclonal antibodies against receptor proteins for estrogen (ER) and progesterone (PR) has opened a new perspective in the endocrinological evaluation of breast cancer patients. Indeed, a great deal of information can be obtained by applying these antibodies to tissue sections, including information that cannot be provided by steroid-binding assays. Of particular interest is the assessment of the percentage of receptor-positive cells in any given tumor, which can be easily evaluated in stained sections, but cannot be estimated by biochemical assays on tissue homogenates. Basic research in this field has been done on frozen sections which have proved to be quite reliable as documented by several reports presented at a recent symposium on Estrogen Receptor Determination with Monoclonal Antibodies (Cancer Res. Suppl. 46: 4231–4314, 1986). However, for the purpose of using immunocytochemistry for the practical assessment of the ER and PR status of breast cancer patients it should be realized that tissue for frozen sections is not always available and for this reason it is important to be able to apply these techniques to paraffin embedded tissues or to cytological preparations.

Initial Clinical Experience with a Holmium:Yag Laser

Transmyocardial laser revascularization (TMLR) has been used in many patients to effectively treat angina that is refractory to medical therapy and which cannot be treated by traditional revascularization techniques. To date, a majority of these procedures have been performed with a carbon dioxide laser. However, because of the potentially large number of patients who could benefit from such a therapy, either as sole therapy or as an adjunct to coronary bypass and angioplasty, several different types of laser energy sources have been developed for TMLR. At least two clinical trials are currently underway investigating the safety and efficacy of holmium: YAG lasers. Our institution has been participating in an international, multicenter study evaluating one such laser; the CardioGenesis ITMR System. As part of this study, fifteen patients have so far undergone TMLR as sole therapy in the United States with the CardioGenesis ITMR System and, as reported in other trials, there has been an immediate and clinically significant reduction in angina observed in most patients [1]. In this chapter, we summarize the rather dramatic results obtained in our first patient in this trial.