Focal angiogen therapy using intramyocardial delivery of an adenovirus vector coding for vascular endothelial growth factor 121.

BACKGROUND: Adenovirus (Ad) vector-mediated gene therapy strategies have emerged as promising modalities for the "biological revascularization" of tissues. We hypothesized that direct intramyocardial, as opposed to intracoronary, administration of an Ad vector coding for the vascular endothelial growth factor 121 cDNA (Ad(GV)VEGF121.10) would provide highly focal Ad genome levels, and increases in VEGF, ideal for inducing localized therapeutic angiogenesis. METHODS: Persistence and regional distribution of the vector were assessed by TaqMan real-time quantitative polymerase chain reaction technology and enzyme-linked immunosorbent assay, after intramyocardial Ad(GV)VEGF121.10 in the rat, and either intramyocardial or intracoronary (circumflex territory) vector in Yorkshire swine. Based on these results, we assessed the focal nature of the improved cardiac blood flow in a previously reported porcine myocardial ischemia model. RESULTS: Intramyocardial delivery of Ad(GV)VEGF121.10 in the rat resulted in local persistence of the Ad genome that decreased 1,000-fold over 3 weeks, with peak myocardial VEGF expression 24 to 72 h after vector delivery. After intramyocardial Ad(GV)VEGF121.10 in the circumflex distribution of pigs, Ad vector genome and VEGF protein levels were more than 1,000-fold and more than 90-fold higher, respectively, in this distribution than in other myocardial regions. In comparison, intracoronary injection yielded maximum myocardial Ad genome and VEGF levels 33-fold and 9-fold lower, respectively, than that after intramyocardial delivery. Angiograms obtained 28 days after intramyocardial Ad(GV)VEGF121.10 demonstrated rapid circumflex reconstitution via collaterals localized to the region of vector administration. CONCLUSIONS: These studies demonstrate that direct intramyocardial administration of Ad(GV)VEGF121.10 results in focal genome and VEGF levels, including focal angiogenesis, sufficient to normalize blood flow to the ischemic myocardium, findings that are relevant to designing human trials of gene therapy-mediated cardiac angiogenesis.

Safety of direct myocardial administration of an adenovirus vector encoding vascular endothelial growth factor 121.

A gene therapy strategy involving direct myocardial administration of an adenovirus (Ad) vector encoding the vascular endothelial growth factor 121 cDNA (Ad(GV)VEGF121.10) has been shown to be capable of "biological revascularization" of ischemic myocardium in an established porcine model [Mack, C.A. (1998). J. Thorac. Cardiovasc. Surg. 115, 168-177]. The present study evaluates the local and systemic safety of this therapy in this porcine ischemia model and in normal mice. Myocardial ischemia was induced in Yorkshire swine with an ameroid constrictor 21 days prior to vector administration. Ad(GV)VEGF121.10 (10(9) or 10(10) PFU), Ad5 wild type (10(9) PFU), AdNull (control vector with no transgene; 10(9) PFU), saline, or no injection (naive) was administered in 10 sites in the ischemic, circumflex distribution of the myocardium. Toxicity was assessed by survival, serial echocardiography, blood analyses, and myocardial and liver histology at 3 and 28 days after vector administration. All pigs survived to sacrifice, except for one animal in the Ad(GV)VEGF121.10 (10(10) PFU) group, which died as a result of oversedation. Echocardiograms of Ad(GV)VEGF121.10-treated pigs demonstrated no differences in pericardial effusion, mitral valve regurgitation, or regional wall motion compared with control pigs. Intramyocardial administration of Ad(GV)VEGF121.10 included only minimal myocardial inflammation and necrosis, and no hepatic inflammation or necrosis. Only a mild elevation of the white blood cell count was encountered on day 3, which was transient and self-limited in the Ad(GV)VEGF121.10 group as compared with the saline-treated animals. As a measure of inadvertent intravascular administration of vector, normal C57/BL6 mice received intravenous Ad(GV)VEGF121.10 (10(4), 10(6), 5 x 10(7), or 10(9) PFU), AdNull (5 x 10(7) or 10(9) PFU), or saline. Toxicity was assessed by survival, blood analyses, and organ histology at 3 and 7 days after vector administration. A separate group of C57/BL6 mice received intravenous AdmVEGF164 (Ad vector encoding the murine VEGF164 cDNA), Ad(GV)VEGF121.10, AdNull (10(8) PFU each group), or saline to assess duration of expression and safety of a homologous transgene. All mice survived to sacrifice except for 40% of the mice in the highest (10(9) PFU; a dose more than 10(3)-fold higher by body weight than the efficacious dose in pigs) Ad(GV)VEGF121.10 dose group, which died on days 5-6 after vector administration. The only differences seen in the blood analyses between treated and control mice were in the very high Ad(GV)VEGF121.10 dose group (10(9) PFU), which demonstrated an anemia as well as an increase in alkaline phosphatase when compared with all other treatment groups. Hepatic VEGF levels by ELISA in AdmVEGF164-treated mice did not persist beyond 14 days after vector administration, suggesting that persistent expression of a homologous VEGF gene transferred with an Ad vector is not a significant safety risk. Although this is not a chronic toxicity study, these data demonstrate the safety of direct myocardial administration of Ad(GV)VEGF121.10, and support the potential use of this strategy to treat human myocardial ischemia.

Salvage angiogenesis induced by adenovirus-mediated gene transfer of vascular endothelial growth factor protects against ischemic vascular occlusion.

PURPOSE: Vascular endothelial growth factor (VEGF) is a potent stimulator of angiogenesis, and transgene expression from adenovirus vectors can provide in vivo delivery of proteins. On the basis of this knowledge, we hypothesized that local administration of a replication-deficient adenovirus vector expressing complementary DNA for VEGF (AdVEGF) would induce collateral vessel formation in the setting of ischemia that could protect against subsequent acute vascular occlusion. METHODS: Hindlimb ischemia was induced in Sprague-Dawley rats by means of unilateral ligation of the common iliac artery immediately followed by administration of 4 x 10(9)-plaque-forming units VEGF, the control vector AdNull, or phosphate-buffered saline solution into the iliofemoral adipose tissue and thigh muscles. Untreated rats with common iliac ligation were used as an additional control group. RESULTS: Local VEGF expression was observed for 5 days in AdVEGF-treated rats but not in controls. Three weeks after ligation and vector administration, the ipsilateral femoral artery was ligated for a model of an acute vascular occlusion in the setting of preexisting ischemia. Blood flow to the ischemic hindlimb relative to the contralateral hindlimb evaluated with color microspheres demonstrated significantly increased blood flow in the AdVEGF-treated rats compared with each control group (p < 0.0001). Relative blood flow assessed by means of 99mTc-sestamibi radionuclide scans also demonstrated increased blood flow to the ligated hindlimb of AdVEGF-treated rats compared with each control group (p < 0.002). AdVEGF-treated rats also demonstrated increased vascularity in the ligated limb compared with each control group as assessed by means of angiography (p < 0.0001) and histologic quantification of blood vessels less than 80 microm diameter in local adipose tissue and capillaries per muscle fiber (p < 0.0002). AdVEGF treatment prevented a rise in femoral venous lactate femoral venous concentrations 1 hour after femoral artery ligation in control rats (p < 0.04). CONCLUSIONS: An adenovirus vector expressing VEGF complementary DNA is capable of stimulating an angiogenic response that protects against acute vascular occlusion in the setting of preexisting ischemia, suggesting that in vivo gene transfer of VEGF complementary DNA might be useful in prophylaxis of advancing arterial occlusive disease.

Biologic bypass with the use of adenovirus-mediated gene transfer of the complementary deoxyribonucleic acid for vascular endothelial growth factor 121 improves myocardial perfusion and function in the ischemic porcine heart.

OBJECTIVES: Vascular endothelial growth factor (VEGF), a potent angiogenic mediator, can be delivered to targeted tissues by means of a replication-deficient adenovirus (Ad) vector. We hypothesized that direct administration of Ad vector expressing the VEGF121 complementary deoxyribonucleic acid (AdGVVEGF121.10) into regions of ischemic myocardium would enhance collateral vessel formation and improve regional perfusion and function. METHODS: Yorkshire swine underwent thoracotomy and placement of an Ameroid constrictor (Research Instruments & MFG, Corvallis, Ore.) on the circumflex coronary artery. Three weeks later, myocardial perfusion and function were assessed by single photon emission computed tomography imaging (SPECT) with 99mTc-labeled sestamibi and by echocardiography during rest and stress. AdGVVEGF121.10 (n = 7) or the control vector, AdNull (n = 8), was administered directly into the myocardium at 10 sites in the circumflex distribution (10(8) pfu/site). Four weeks later, these studies were repeated and ex vivo angiography was performed. RESULTS: SPECT imaging 4 weeks after vector administration demonstrated significant reduction in the ischemic area at stress in AdGVVEFG121.10-treated animals compared with AdNull control animals (p = 0.005). Stress echocardiography at the same time demonstrated improved segmental wall thickening in AdGVVEGF121.10 animals compared with AdNull control animals (p = 0.03), with AdGVVEGF121.10 animals showing nearly normalized function in the circumflex distribution. Collateral vessel development assessed by angiography was also significantly greater in AdGVVEGF121.10 animals than in AdNull control animals (p = 0.04), with almost complete reconstitution of circumflex filling in AdGVVEGF121.10 animals. CONCLUSIONS: An Ad vector expressing the VEGF121 cDNA induces collateral vessel development in ischemic myocardium and results in significant improvement in both myocardial perfusion and function. Such a strategy may be useful in patients with ischemic heart disease in whom complete revascularization is not possible.

Channel patency and neovascularization after transmyocardial revascularization using an excimer laser: results and comparisons to nonlased channels.

BACKGROUND: Transmyocardial revascularization (TMR) has emerged as a promising treatment for ischemic heart disease in patients who are not candidates for coronary bypass surgery or angioplasty. Controversy exists, however, as to whether the use of laser energy is critical for TMR channel patency. We therefore compared by histologic assessment the outcome of lased channels with nonlased channels 30) days after TMR, using a low energy, short-pulse, fiberoptic excimer laser. METHODS AND RESULTS: In each of six sheep, 36 1-mm TMR channels (9 mJ; 240 Hz; 1.55 cm advance/s) were placed in the anterior wall of the left ventricle, and 12 1-mm nonlased channels were created in adjacent segments by advancing the fiberoptic through the left ventricular wall with the laser inactivated. Of the 36 lased channels, 56+/-7.3% were identifiable, and 100% of those identifiable appeared to represent a "channel derivative" with evidence of an endothelialized lumen, whereas none of the nonlased channels had evidence of channel patency. Lased channels had a marked neovascular response (graded on a 0-3 scale) compared with nonlased channels (2.5+/-0.1 versus 1.0+/-0.1; P<.05). Echocardiography performed 1 and 30 days after TMR demonstrated normal global and regional left ventricular function in all animals. Creatinine phosphokinase myocardial band fractions were not significantly increased after TMR. CONCLUSIONS: TMR using the excimer laser results in increased evidence of channel derivatives and neovascularization compared with nonlased channels while preserving normal ventricular function. These findings suggest that laser energy may be an important component of TMR strategy.

Therapeutic angiogenesis: a comparative study of the angiogenic potential of acidic fibroblast growth factor and heparin.

PURPOSE: Acidic fibroblast growth factor (aFGF) is a potent mitogen for vascular and other mesenchymal cells in vitro that can induce angiogenesis in vivo. Although heparin has no mitogenic potential of its own, it is an important aFGF cofactor in vitro and may also be capable of stimulating angiogenesis. Because the development of a collateral vasculature in response to ischemia appears to be dependent on angiogenesis, we compared the abilities of aFGF with or without heparin and heparin alone to accelerate angiogenesis in a rat hind limb ischemia model. METHODS: Daily subcutaneous injections of saline solution (1 ml), heparin (0.05 mg), or human recombinant aFGF with or without heparin (1 microgram aFGF, 0.05 mg heparin) were administered into the hind limb region distal to the point of unilateral femoral artery ligation in the rat for the 10 days immediately after vascular occlusion. Angiogenicity was determined by histologic assessment of treatment outcomes. RESULTS: Histologic assessment of the number of vessels per microscopic field 10 days after vascular ligation in the fibrofatty tissues distal to the ligation point had the following results: saline solution, 10 +/- 4 vessels; heparin, 13 +/- 4 vessels (p < 0.05 vs saline solution); aFGF, 26 +/- 8 vessels; and aFGF/heparin 36 +/- 8 vessels (aFGF, aFGF/ heparin, p < 0.001 vs saline solution). Similar increases in vascularization were also noted in the skeletal muscle tissues distal to the vascular ligation point. Immunohistochemical analysis for the presence of proliferating cell nuclear antigen, a marker for mitogenic activity, demonstrated corresponding increases in proliferating cell nuclear antigen labeling for each of the treatment groups, expressed as a percentage of total vascular cell nuclei, as follows: saline solution, 7% +/- 2%; heparin, 21% +/- 8% (p < 0.05 vs saline solution); aFGF, 67% +/- 9%; and aFGF/heparin, 83% +/- 5% (aFGF, aFGF/heparin, p < 0.001 vs saline solution). CONCLUSIONS: The increased vascularization and mitogenic activity demonstrated by these respective studies suggest that angiogenesis is significantly accelerated by the administration of heparin alone and is accelerated to a greater extent by the administration of aFGF with or without heparin. The aFGF/heparin regimen may represent an optimal means of augmenting collateral vessel growth to relieve ischemia in the clinical setting.

Augmentation of blood platelet levels by intratracheal administration of an adenovirus vector encoding human thrombopoietin cDNA.

This study was designed to evaluate the hypothesis that administration of a replication-deficient, recombinant adenovirus vector to the epithelial surface of the respiratory tract can be used to deliver a recombinant protein to the systemic circulation in sufficient quantities to evoke a systemic response appropriate to the recombinant protein. We administered AdCMV.TPO-an adenovirus vector containing an expression cassette coding for the human thrombopoietin (TPO) cDNA-to the respiratory epithelium of immunocompetent Balb/c mice. Over the following week, serum human TPO levels were elevated, platelet levels increased more than sixfold, and megakaryocytosis was evident in bone marrow. This strategy may be a useful approach to the nonparenteral administration of a variety of therapeutic recombinant proteins, such as those relevant to clotting, endocrine function, and bone-marrow function.

Vascular endothelial growth factor is the major angiogenic factor in omentum: mechanism of the omentum-mediated angiogenesis.

Omentum has been used clinically to promote wound healing and to stimulate the revascularization of ischemic tissues. The biologic mechanism responsible for these effects has, however, not yet been defined. A number of polypeptide growth factors that possess potent angiogenic properties have recently been identified, and we therefore sought to determine whether one of these growth factors might be responsible for the angiogenic properties of the omentum. The levels of vascular endothelial growth factor (VEGF) protein in a number of rat tissues and organs were analyzed by Western and enzyme immunoassay analysis. Because omentum was found to have the greatest VEGF concentrations of the tissues examined, antibody neutralization, transcription inhibition assays, and Northern blot analysis were performed under hypoxic and normoxic conditions on tissues extractions and primary tissue cultures of omentum to further characterize the functional significance of VEGF expression in these tissues. The omentum demonstrated the highest VEGF secretion rate as well as the highest concentration of VEGF protein of the various rat tissues and organs examined. Fractionation studies of the omentum furthermore demonstrated that omental adipocytes, rather than the stromal-vascular cells, were the primary source of VEGF protein. An endothelial cell mitogenic assay showed that a major portion of the mitogenic activity of heparin-binding proteins and conditioned media derived from omentum was abolished by VEGF antibody. Additional studies with the transcription inhibitor actinomycin-D furthermore demonstrated that the VEGF gene was continuously transcribed in the rat omental adipocytes. Incubation of the omental adipocytes under hypoxic conditions induced approximately a 1.7-fold increase in VEGF protein expression, which was abolished by actinomycin-D. Northern blot analysis demonstrated that hypoxia resulted in upregulation of the VEGF mRNA in the hypoxia-cultured omental adipocytes, suggesting that the augmentation of VEGF expression in omental adipocytes by hypoxia occurs at the transcriptional level. These data suggest that VEGF is the major angiogenic factor produced by omentum and possibly underlies the mechanism of omentum-induced angiogenesis. Augmented expression of VEGF by omental cells under hypoxic conditions may furthermore reflect the mechanism responsible for enhancing the angiogenic activity of omentum in the setting of ischemia.

Regional angiogenesis induced in nonischemic tissue by an adenoviral vector expressing vascular endothelial growth factor.

The feasibility of a single administration of a replication-deficient adenovirus (Ad) vector encoding the cDNA for human vascular endothelial growth factor (VEGF) (AdCMV.VEGF) to induce neovascularization in vivo in normal tissue was evaluated in retroperitoneal adipose tissue. Following administration of AdCMV.VEGF (10(9) pfu/50 microliters), maximal VEGF cDNA expression was observed at 2-5 days in the injected adipose tissue. No VEGF protein was detected at > or = 10 days in injected adipose tissue, and there was no increase in serum VEGF levels at any time. In vivo quantification of the number of blood vessels using 30x visualization of the adipose tissue demonstrated an increase in vessel number by 10 days, plateauing by 30 days with a 123% increase in vessel number compared to the control vector AdCMV.Null, despite the fact that no VEGF protein was detected after 5 days. Consistent with the in vivo data, histologic quantification of capillary number demonstrated an increase by day 5, reaching a 38% increase over AdCMV.Null by day 30. These observations demonstrate that an Ad vector carrying the VEGF cDNA is capable of inducing the growth of new blood vessels in a regional fashion in a relatively avascular, normal organ. This suggests in vivo Ad-mediated gene transfer may be useful for therapeutic angiogenesis in the treatment of ischemic cardiovascular disease.

Circumvention of anti-adenovirus neutralizing immunity by administration of an adenoviral vector of an alternate serotype.

Effective gene transfer and expression following repetitive administration of adenoviral (Ad) vectors in experimental animals is limited by anti-Ad neutralizing antibodies. Knowing that anti-Ad humoral immunity is serotype-specific, we hypothesized that anti-Ad neutralizing immunity could be circumvented using Ad vectors of different serotypes (Ad2, Ad5) within the same subgroup (C) to transfer and express beta-glucuronidase (beta glu) in the lung. Sprague-Dawley rats received an intratracheal administration of either Ad2 beta glu or Ad5 beta glu, and, 14 days later, repeat administration of either the same vector or a vector of a different serotype. Analysis of serum and bronchoalveolar lavage fluid following initial vector administration demonstrated systemic and local serotype-specific neutralizing antibodies. For both the Ad2 and Ad5 vectors, beta glu expression 24 hr following the second administration of the same serotype was < 30% of that of naive animals. In contrast, beta glu expression 24 hr following second administration of a different serotype Ad vector was similar to expression at 24 hr of naive animals receiving a single administration (Ad5 beta glu followed by Ad2 beta glu, as well as Ad2 beta glu followed by Ad5 beta glu; p > 0.2 both comparisons). Although the alternative serotype bypassed anti-Ad neutralizing immunity, persistence of expression was reduced compared to that following administration to naive animals. Compatible with this observation, systemic administration of the same vectors to C57B1/6 mice demonstrated induction of cytotoxic T lymphocytes directed against the beta glu transgene, as well as products of the Ad genome. Interestingly, intratracheal administration of vectors with different serotypes and different transgenes to rats resulted in longer expression (but still not normalized) compared to that achieved with vectors of different serotypes but the same transgene. These observations demonstrate that alternate use of Ad vectors from different serotypes within the same subgroup can circumvent anti-Ad humoral immunity to permit effective gene transfer after repeat administration, although the chronicity of expression is limited, likely by cellular immune process directed against both the transgene and viral gene products expressed by the vector.

Myocardial angiogenesis as a possible mechanism for TMLR efficacy.

Despite advances in the treatment of ischemic heart disease, there still exists a significant number of individuals for whom bypass surgery or angioplasty are not options. Transmyocardial laser revascularization (TMLR) is a promising technology that has already been shown to reduce symptoms in patients with chronic ischemic heart disease that is not amenable to conventional therapies. Although it appears that TMLR can provide symptomatic relief of angina in selected patients, the mechanism by which TMLR is thought to work is unclear. Recently it has been postulated that TMLR induces an angiogenic response and, perhaps, improves local perfusion to ischemic myocardial territories. A brief overview of the biology of myocardial angiogenesis is presented.

Primary inflammatory fibrosarcoma of the esophagus.

Primary sarcomas of the esophagus are rare. We report the radiologic, surgical, and pathologic findings of a primary inflammatory fibrosarcoma of the esophagus in a 33-year-old woman, and review the prognostic features and management options of this tumor.

Direct in vivo gene transfer to canine myocardium using a replication-deficient adenovirus vector.

BACKGROUND: Direct myocardial gene transfer is a mordality that involves the introduction of genetic information into myocardial tissue to achieve a therapeutic effect. This study was designed to characterize the temporal and spatial limits of gene expression and to determine the safety of direct myocardial gene transfer in a large animal model using replication-deficient adenovirus vectors. METHODS: Mongrel dogs underwent left thoracotomy and direct myocardial injections (100 microL/injection) of adenovirus vectors (10(9) pfu) carrying the DNA for the reporter enzyme chloramphenicol acetyl transferase or the angiogenic protein vascular endothelial growth factor. Two to 14 days after vector administration, regional protein expression was evaluated in myocardium and distant organs. Left ventricular function, assessed by echocardiography, and routine hematologic and biochemical indices were evaluated before and after vector administration. RESULTS: Peak levels of chloramphenicol acetyl transferase activity were detected 2 days after vector administration, and levels above baseline persisted for at least 14 days. Local chloramphenicol acetyl transferase activity was detected at distances at least as far as 1.5 cm from the site of injection. Chloramphenicol acetyl transferase activity in distant organs was less than 0.1% of that in injected myocardium 7 days after vector administration. Localized expression of vascular endothelial growth factor was achieved for up to 7 days after a single vector administration. Cardiac function and laboratory values were unchanged during the study. CONCLUSIONS: Adenovirus-mediated direct myocardial gene transfer can be accomplished safely in a large animal model, providing high levels of protein expression in a greater spatial distribution than previously reported, with minimal transfection of distant organs. Sustained and localized expression of a potent angiogenic mediator has been accomplished, which may provide an innovative strategy to stimulate angiogenesis in ischemic myocardium.

A technique for laparoscopic repair of herniation of the anterior abdominal wall using a composite mesh prosthesis.

Improved laparoscopic techniques have engendered many new gastrointestinal and other intracavity abdominal procedures. Groin hernias have also been repaired with the assistance of the laparoscope via both transperitoneal and properitoneal approaches, but less emphasis has been placed upon repair of hernias of the anterior abdominal wall. A technique for the transperitoneal, laparoscopic repair of anterior abdominal wall hernias using a composite mesh prosthesis is presented. The technique is applicable to hernias in many locations.

Planned laparoscopic repair of a spigelian hernia using a composite prosthesis.

A planned elective repair, via the laparoscope, of a spigelian hernia is described. The repair was performed using a composite mesh prosthesis consisting of a sandwich of polyester fiber mesh and polyglactin 910 mesh, sutured together with polyglactin 910 suture at the operating table before introduction. The technique is applicable to other hernias of the anterior abdominal wall.

Clinical and hemodynamic performance of the 19-mm Carpentier-Edwards porcine bioprosthesis.

Because of concerns about the hemodynamic performance of 19-mm porcine valves, we retrospectively reviewed the clinical results and echocardiographic studies of 52 consecutive patients who received a 19-mm Carpentier-Edwards porcine bioprosthesis (model 2625) for aortic valve replacement from 1986 through 1991. Nearly 87% of the patients were women, the mean age was 69 years, and the mean body surface area was 1.63 +/- 0.27 m2. Seventy-three percent of the patients had pure aortic stenosis, 96% were in New York Heart Association classes III and IV, and 56% underwent urgent or emergent operation. Overall hospital mortality was 7.7% with a late mortality of 8.3% at a mean follow-up of 25 +/- 18 months. No patient experienced a valve-related complication, and 95% of surviving patients were in New York Heart Association classes I and II. Two-dimensional and Doppler echocardiography performed during the first postoperative week revealed a maximal instantaneous gradient of 44.7 +/- 13.0 mm Hg. In 43 patients for whom additional data were available, the mean gradient was 26.4 +/- 8.2 mm Hg with an effective orifice area of 0.85 +/- 0.18 cm2. This study defines the normal range of postoperative gradients across the 19-mm Carpentier-Edwards porcine valve and demonstrates that patients receiving this valve can achieve significant clinical improvement despite the presence of high transvalvular gradients measured by echocardiography.

Absorption and exposure in positive photoresist.

A review of the theory of absorption on microscopic and macroscopic levels is given. This theory is then applied to the absorption of UV light by diazo-type positive photoresist during exposure. A formal treatment of the properties of polychromatic light is given. Using these analyses, the effects of polychromatic exposure of a photoresist are derived. Finally, experimental verification of Beer's law and determination of the exposure quantum efficiency of a particular photoresist is given.

Myogenic vasoregulation overrides local metabolic control in resting rat skeletal muscle.

Microvascular reactions to increases in intravascular pressure were studied in the cremaster muscle of the anesthetized rat by enclosing the animal in an airtight box with the muscle exteriorized for observation of the microcirculation. Since the cremaster was exposed to atmospheric pressure, increasing pressure within the box produced equal increases in arterial and venous pressures. Thus, intravascular pressure was altered without affecting the pressure gradient for blood flow. Raising box pressure had no effect on respiration or heart rate and did not change the systemic activity of the sympathetic system, angiotensin II, or vasopressin. Diameters and flows were measured for first (107 +/- 3 micron, mean +/- SEM), second (87 +/- 5), third (29 +/- 2), and fourth (15 +/- 2) order arterioles during increases in intravascular pressure of +10, +20, and +30 mm Hg. No significant changes in the diameters of first or second order arterioles were elicited when pressure was increased. However, when box pressure was increased to +10, +20, or +30 mm Hg, a sustained constriction occurred in third (29%, 45%, and 63%, respectively) and fourth (5%, 38%, and 57%, respectively) order arterioles. Blood flow was significantly reduced in all arterioles, and perivascular PO2 was decreased adjacent to third and fourth order arterioles. Furthermore, the third order arteriole constrictor response was not abolished by local alpha-receptor blockade (phentolamine), indicating that it was not mediated by a local sympathetic axon reflex. Collectively, these data indicate that a potent, non-neural, pressure-dependent mechanism for vasoregulation is present in small arterioles of the cremaster. The sustained constriction in the presence of reduced blood flow and reduced periarteriolar oxygen tension indicates that the vascular response is independent of and capable of overriding flow-dependent (i.e., metabolic) control in resting skeletal muscle. The observations are compatible with the operation of a powerful myogenic mechanism in small arterioles.