Predicting Factors Associated with Postoperative Hypotension following Carotid Artery Stenting.

BACKGROUND: Prolonged hemodynamic instability after carotid artery stenting (CAS) has been associated with increased incidence of stroke and other major adverse events. The objective of this study is to determine the factors associated with hypotension following CAS. In particular, this study evaluates whether involvement of the carotid bifurcation/bulb and degree of calcification can predict postoperative hypotension. METHODS: A retrospective review of 90 CASs performed in 88 patients at a single tertiary center was completed. In patients with proximal internal carotid stenosis involving the carotid bifurcation, the extent of bifurcation/bulb calcification on preoperative computed tomography angiography was assessed using a scoring system. Calcium scores were assigned based on the percent of circumferential calcification of carotid bifurcation as follows: grade 1, <10%; grade 2, 10-50%; grade 3, 50-90%; and grade 4, >90%. Perioperative factors associated with prolonged postoperative hypotension requiring vasopressor infusion were analyzed. RESULTS: Overall, postoperative hypotension requiring vasopressors occurred in 26 (28.9%) of CAS. There were no differences in baseline demographics, comorbidities, or CAS indication between patients who required postoperative vasopressors for hypotension and those who did not. The majority of patients (64.4%) were on 2 or more antihypertensive medications preoperatively. Stenosis involved carotid bifurcation in 64 (71.1%) cases. Of these, 27 (42.2%) were grade 1, 19 (29.7%) were grade 2, 10 (15.6%) were grade 3, and 8 (12.5%) were grade 4 based on our calcium scoring system. On risk-adjusted analysis, carotid bifurcation/bulb involvement (adjusted odds ratio [aOR] 4.5, 95% confidence interval [CI] 1.1-18.5) and preoperative regimen of 2 or more antihypertensives (aOR 4.2, 95% CI 1.1-16.0) were independent predictors of hypotension requiring vasopressors following CAS. Among patients with carotid bifurcation involvement, severity of calcium score was not a significant predictor of postoperative hypotension. CONCLUSIONS: CAS for carotid stenosis involving the carotid bifurcation/bulb is associated with a higher risk for postoperative hypotension requiring vasopressors. Patients with preoperative hypertension requiring 2 or more antihypertensive medications are also at increased risk. However, severity of carotid bifurcation calcification is not a significant predictor of need for postoperative vasopressors.

Hybrid Revascularization Combining Iliofemoral Endarterectomy and Iliac Stent Grafting for TransAtlantic Inter-Society Consensus C and D Aortoiliac Occlusive Disease.

BACKGROUND: This study examines the outcome of hybrid revascularization combining iliofemoral endarterectomy and iliac artery stenting using covered stents in TransAtlantic Inter-Society Consensus (TASC) C and D aortoiliac occlusive disease (AIOD) involving the common femoral artery (CFA). METHODS: A retrospective review was conducted in patients with TASC C and D AIOD involving the CFA and undergoing hybrid revascularization. Covered stents were used primarily. Demographics, indications for surgery, lesion classification, hospital length of stay (LOS), 30-day morbidity/mortality, hemodynamic and clinical success, and patency were assessed. RESULTS: Thirty-six male patients (41 limbs), mean age 63.9 ± 6 years, were identified (TASC C = 39%, D = 61%). Indications for surgery were claudication (27%), rest pain (44%), and tissue loss (29%). A simultaneous adjunctive procedure (5 infrainguinal bypass, 3 superficial femoral artery stents) was performed in 22%. Thirty-day outcomes included 1 mortality (2.7%) and 2 reoperation (5.5%), 1 for femoral artery pseudoaneurysm and 1 for bilateral groin seroma. LOS was 4 days (interquartile range 3-6). All patients with available data experienced 30-day clinical and hemodynamic success. Mean follow-up was 23 months (range 1-79 months) with a primary patency of 85.4%. Cumulative primary assisted and secondary patency was 92.6%. The femoral patch repair was the most frequent site of reintervention (3/3). Mortality was 34% during the study period, and it was significantly higher in patients with tissue loss (57.1% vs. 14.8%, P = 0.01). CONCLUSIONS: The hybrid approach has low morbidity, mortality, and fast recovery. The use of covered stents/stent grafts provides good mid-term patency. Close follow-up with noninvasive imaging is paramount to avoid repair failure, in particular at the femoral patch repair site.

Endovascular treatment of TransAtlantic Inter-Society Consensus D aortoiliac occlusive disease using unibody bifurcated endografts.

OBJECTIVE: Aortobifemoral bypass has been the gold standard treatment for extensive aortoiliac occlusive disease. Endovascular therapy and stenting of aortic and iliac occlusive lesions has proven to be efficacious, especially when dealing with short segment lesions. Endovascular treatment of TransAtlantic Inter-Society Consensus II (TASC) D aortoiliac occlusive lesions remains a challenge, but a valuable treatment option in poor surgical candidates. We present our operative technique and midterm results in treating TASC D aortoiliac occlusive disease using unibody bifurcated endografts. METHODS: We performed a retrospective review of patients with TASC D aortoiliac occlusive disease who underwent transfemoral endovascular revascularization with the Endologix Powerlink unibody bifurcated endograft (Endologix, Irvine, Calif). Demographic data, operative details, and outcomes were collected. Paired t-tests were performed to compare preoperative and postoperative ankle brachial indexes. RESULTS: Between March 2009 and July 2011, 10 high-risk patients (8 male and 2 female) for a traditional aortobifemoral bypass were treated using this endovascular technique. The mean age was 59 ± 6 years (range, 50-69 years). All patients presented with rest pain, and four with tissue loss. Technical success was 100%, with two patients requiring brachial access and eight patients requiring additional stent placement. Postoperatively, all patients reported clinical improvement with resolution of ischemic symptoms. Mean improvement ankle brachial index was 0.50 ± 0.08 (P = .028) and 0.50 ± 0.01 (P = .034) in the left and right legs, respectively. Mean follow-up time was 40 ± 24 months (range, 4-81 months). The primary and secondary patency rates were 80% and 100%, respectively. Complications requiring early reintervention occurred in two patients and included one expanding hematoma from the percutaneous access site and one acute iliac artery thrombosis. Additionally, one patient underwent repeat angioplasty/stenting for threatened endograft limbs at 4 months. One patient expired during follow-up from an unrelated cardiac cause 19 weeks postoperatively. CONCLUSIONS: This series demonstrates that endovascular repair using a unibody bifurcated endograft for TASC D aortoiliac occlusive disease is feasible, effective, and has excellent midterm patency. It should be considered an effective treatment option when the disease process involves the aorta, in particular if the patient is surgically unfit for a traditional aortobifemoral bypass. The unibody configuration preserves the anatomic aortic bifurcation, which is particularly important in patients with peripheral occlusive disease who are deemed to undergo subsequent endovascular interventions.

Extra-anatomic endovascular repair of an abdominal aortic aneurysm with a horseshoe kidney supplied by the aneurysmal aorta.

Abdominal aortic aneurysm complicated by a horseshoe kidney (HSK, fused kidney) represents a unique challenge for repair. Renal arteries arising from the aneurysmal aorta can further complicate intervention. Reports exist describing the repair of these complex anatomies using fenestrated endografts, hybrid open repairs (debranching), and open aneurysmorrhaphy with preservation of renal circulation. We describe an extra-anatomic, fully endovascular repair of an abdominal aortic aneurysm with a HSK partially supplied by a renal artery arising from the aneurysm. We successfully applied aortouni-iliac endografting, femorofemoral bypass, and retrograde renal artery perfusion via the contralateral femoral artery to exclude the abdominal aortic aneurysm and preserve circulation to the HSK.

"Cheese wire" fenestration of a chronic aortic dissection flap for endovascular repair of a contained aneurysm rupture.

A 70-year-old man presented with a contained aortic rupture above the aortic bifurcation in the setting of a chronic type B aortic dissection. The celiac and right iliac arteries were supplied by the false lumen, and the superior mesenteric, left iliac, and bilateral renal arteries were supplied by the true lumen. An open repair was not possible due to right-sided heart failure. The "cheese wire" maneuver is a technique used to fenestrate an intimal flap, alleviating malperfusion in aortic dissection. In our modification, a Glidewire (Terumo Medical, Somerset, NJ) was passed across the intimal flap using a Ross Modified Colapinto needle (Cook Medical, Bloomington, Ind) and pulled downward to shear through the membrane to the aortic bifurcation. The fenestration was followed by a unibody endograft stent repair across the contained rupture.

Hyperbaric oxygen for chronic wounds.

Hyperbaric oxygen therapy (HBOT), the administration of pressurized 100% oxygen, is used as an adjunct to aid healing in selected chronic wounds. Though the therapy has had a controversial history, research is now elucidating the mechanisms by which HBOT helps to heal wounds. HBOT increases growth factors and local wound signaling, while also promoting a central stem cell release of endothelial progenitor cells from the bone marrow via nitric oxide pathways. The clinical data continue to accumulate in support of HBOT to help hasten wound healing, and reduce the amputation rate in diabetic ulcers. In appropriate patients, HBOT is an effective, noninvasive, adjunct modality that can be used to hasten chronic wound healing.

Modified use of the Hemodialysis Reliable Outflow (HeRO) graft for salvage of threatened dialysis access.

The Hemodialysis Reliable Outflow (HeRO) graft (Hemosphere Inc, Eden Prairie, Minn) offers a new option to provide upper extremity arteriovenous (AV) dialysis access in patients with central venous occlusive disease. Creative use of this device can allow for salvage of failing or threatened AV fistulas and grafts. We present two patients who underwent a modified implantation of the HeRO device for immediate salvage of a malfunctioning AV access. Ipsilateral central venous occlusions were successfully overcome by anastomosing a HeRO device to the existing AV access and tunneled across the chest to the contralateral internal jugular vein.

The use of gadolinium for arterial interventions.

BACKGROUND: Gadolinium (Gd) has been traditionally used as a non-nephrotoxic alternative to iodinated contrast for digital subtraction angiography (DSA) in patients with chronic renal insufficiency. However, its use has been questioned on the basis of reports of nephrotoxicity and its recent association with nephrogenic systemic fibrosis (NSF), a potentially lethal complication. Recently available data are conflicting with respect to the true safety profile of intra-arterial Gd. The purpose of this study was to examine the risk of contrast nephropathy and NSF after Gd exposure in a large population of azotemic patients undergoing DSA. METHODS: A comprehensive database encompassing data on all patients who underwent DSA between June 2003 and December 2007 at the New York Presbyterian Hospital was retrospectively reviewed. Patients receiving Gd either alone or in combination with iodinated contrast during DSA were identified and further analyzed. Acute renal failure (ARF) was defined as an elevation in serum creatinine (Cr) by >0.5 mg/dL within 48 hours of exposure. Clinical follow-up was conducted through chart reviewing as well as telephonic interviews with patients and their primary care physicians. RESULTS: A total of 153 patients underwent 179 exposures to Gd either alone (33%) or in combination (67%) with iodinated contrast. Mean follow-up duration was 27.1 months. The mean Cr level was 1.94 ± 0.78 mg/dL and 1.96 ± 1.1 mg/dL before and after DSA, respectively. There were 20 (11.2%) instances of ARF. The mean Cr level before DSA was higher in patients who developed ARF versus those in the non-ARF group (2.7 ± 1.1 mg/dL vs. 1.9 ± 0.7 mg/dL, p = 0.004). In the ARF group, 12 patients had a return to baseline renal function, four experienced irreversible renal deterioration, and four needed dialysis (4.5% incidence of irreversible renal failure). There were 19 deaths at the time of this study (12.4%). The highest risk for the development of ARF after Gd exposure occurred in patients with Cr levels of >3.0 mg/dL before DSA and in those receiving >0.4 mmol/kg of Gd. For patients who received iodinated contrast in combination with Gd, there was a trend toward a higher risk for developing ARF as compared with those receiving only Gd. Finally, there were no instances of NSF identified in any of the patients who received intra-arterial Gd. CONCLUSIONS: Although Gd has the potential to cause kidney injury similar to iodinated contrast, the risk of irreversible renal failure and the requirement for dialysis is low. Life- or limb-threatening interventions should not be avoided in this patient cohort because of preexisting elevations in Cr. These data should help guide the use of Gd in patients with chronic renal insufficiency.

Endovascular aneurysm repair in nonagenarians is safe and effective.

OBJECTIVES: Advanced age is a significant risk factor that has traditionally steered patients away from open aneurysm repair and toward expectant management. Today, however, the reduced morbidity and mortality of aortic stent grafting has created a new opportunity for aneurysm repair in patients previously considered too high a risk for open surgery. Here we report our experience with endovascular aneurysm repair (EVAR) in nonagenarians. METHODS: Retrospective chart review identified all patients>90-years-old undergoing EVAR over a 9-year period at our institution. Collected data included preoperative comorbidities, perioperative complications, endoleaks, reinterventions, and long-term survival. RESULTS: 24 patients underwent EVAR. The mean age was 91.5 years (range 90-94) among 15 (63%) males and 9 (37%) females. Mean abdominal aortic aneurysm diameter was 6.3±1.1 cm. Eight patients (33%) were symptomatic (pain or tenderness). There were no ruptures. Fourteen patients (58%) had general anesthesia while 10 (42%) had local or regional anesthesia. Mean postoperative length of stay was 3.2±2.4 days (2.8±1.9 days for asymptomatic vs 4.1±3.2 days for symptomatic, P=.29). There was one perioperative mortality (4.2%). There were two local groin seromas (8.3%) and six systemic complications (25%). One patient required reintervention for endoleak (4.2%). There were no aneurysm related deaths beyond the 30-day postoperative period. Mean survival beyond 30 days was 29.7±18.0 months for patients expiring during follow-up. Cumulative estimated 12, 24, and 36-month survival rates were 83%, 64%, and 50%, respectively. Linear regression analysis demonstrated an inverse relationship between the number of preoperative comorbidities and postoperative survival in our cohort (R2=0.701), with significantly decreased survival noted for patients presenting with >5 comorbidities. Those still alive in follow-up have a mean survival of 36.1±16.0 months. CONCLUSION: This is the largest reported EVAR series in nonagenarians. Despite their advanced age, these patients benefit from EVAR with low morbidity, low mortality, and mean survival exceeding 2.4 years. Survival appears best in those patients with ≤5 comorbidities. With or without symptoms, patients over the age of 90 should be considered for EVAR.

Delayed permanent paraplegia after endovascular repair of abdominal aortic aneurysm.

Spinal cord ischemia is a rare complication after abdominal aortic surgery and has been attributed to surgical devascularization of the spinal cord, atheroembolization of the cord circulation, or hypoperfusion of cord structures secondary to hypotension or cord edema. We present a diabetic, hypertensive 75-year-old male with endstage renal disease who presented with a 5.5 cm asymptomatic infrarenal abdominal aortic aneurysm, and concomitant 3.5 cm right common iliac artery aneurysm. After undergoing successful endovascular repair with an aorto-uni-iliac device, unilateral hypogastric artery embolization, and femoral-femoral bypass, he was discharged to a rehabilitation facility neurologically intact with a stage 2 decubitus ulcer. He returned on postoperative day 21 with a large stage 4 septic decubitus ulcer, fever, leukocytosis, hypotension, and paraplegia. We hypothesize that the compromised blood flow from the initial reconstruction, combined with the delayed hypotension imposed by sepsis, resulted in spinal cord infarction. He was eventually discharged to a nursing facility with no improvement in his neurologic status. We report the first case of significantly delayed permanent paraplegia after endovascular abdominal aortic aneurysmorrhaphy.

Successful revascularization for delayed presentation of radiation-induced distal upper extremity ischemia.

Despite several reports of proximal arm ischemia due to radiation therapy, there are no reports of hand ischemia, presumably due to the rarity of radiation treatment of the distal upper extremity. We present a case of a 42-year-old male presenting with acute hand ischemia 36 years after being treated with forearm radiation for Ewing's sarcoma. Angiography demonstrated a patent brachial artery, occluded radial and ulnar arteries in the forearm, and a normal-caliber reconstituted radial artery at the anatomical snuffbox feeding a patent palmar arch. Transluminal balloon angioplasty was attempted initially without improvement. The patient was successfully revascularized with a reversed saphenous vein graft bypass from the distal brachial artery to the distal radial artery. At 22 months of follow-up, the graft remains patent with a palpable distal pulse. The patient continues to report acceptable function and range of motion.

Carotid artery stenting is safe and associated with comparable outcomes in men and women.

OBJECTIVE: Historically, large randomized controlled studies looking at carotid endarterectomy (CEA) have indicated an increased perioperative risk for women when gender subgroup analysis was performed. However, the outcomes of carotid stenting in women as compared to men have not been adequately investigated. We sought to compare the safety and efficacy of carotid angioplasty and stenting (CAS) when performed in women as compared to men. METHODS: Procedures, complications, demographics, co-morbidities, and follow-up data from carotid stenting procedures performed in a bi-campus division were entered into a prospective database and then retrospectively supplemented with stored angiographic image data and reviewed. Arterial anatomic characteristics evaluated using angiographic images were: common carotid/internal carotid lesion length ratio, common carotid/internal carotid diameter, index lesion length, common carotid/internal carotid artery tortuosity, and lesion and aortic arch calcification. Outcomes compared included groin complications, postoperative pressor requirements, length of stay, restenosis, stroke, myocardial infarction (MI), and death. RESULTS: Between 2003 and 2008, 228 patients underwent 238 procedures. Cerebral protection devices and self-expanding stents were placed in all patients. A total of 97 percutaneous interventions performed in 93 women were compared with 141 interventions in 135 men. Mean age in women was 71.8 +/- 9.2 years, in men was 72.2 +/- 9.1 years (P > .99); 44.3% of women and 34.7% of men had symptomatic disease (P = .14). Preoperative demographics and co-morbidities did not differ significantly between genders, with the exception of hypertension (83.0% of males vs 96.7% of females, P = .001), and history of coronary artery bypass grafting (31.8% of males vs 16.1% of females, P = .01). There were no significant differences seen in anatomic arterial characteristics, though there was a trend towards women having larger internal carotid to common carotid diameter ratios (0.65 vs 0.62) and more plaques isolated to the common carotid segment (9.5% vs 6.9%). There were no significant differences seen in overall 30-day peri-procedural stroke rate (2.1% in women and 4.2% in men, P = .48), death rate (0 % vs 0.7%, P > .99), or cardiac events (3.2% vs 0.7%, P = .3). The combined 30-day stroke, death, and MI rate was 5.7% for males compared to 5.4% for females (P > .99). There were no differences observed in the long-term survival, stroke-free survival, or restenosis between genders. CONCLUSION: Despite previous concerns over adverse outcomes in women undergoing carotid endarterectomy, from our data, carotid stenting appears to be a safe modality in women with equivalent outcomes when compared to men.

Diabetic impairments in NO-mediated endothelial progenitor cell mobilization and homing are reversed by hyperoxia and SDF-1 alpha.

Endothelial progenitor cells (EPCs) are essential in vasculogenesis and wound healing, but their circulating and wound level numbers are decreased in diabetes. This study aimed to determine mechanisms responsible for the diabetic defect in circulating and wound EPCs. Since mobilization of BM EPCs occurs via eNOS activation, we hypothesized that eNOS activation is impaired in diabetes, which results in reduced EPC mobilization. Since hyperoxia activates NOS in other tissues, we investigated whether hyperoxia restores EPC mobilization in diabetic mice through BM NOS activation. Additionally, we studied the hypothesis that impaired EPC homing in diabetes is due to decreased wound level stromal cell-derived factor-1alpha (SDF-1alpha), a chemokine that mediates EPC recruitment in ischemia. Diabetic mice showed impaired phosphorylation of BM eNOS, decreased circulating EPCs, and diminished SDF-1alpha expression in cutaneous wounds. Hyperoxia increased BM NO and circulating EPCs, effects inhibited by the NOS inhibitor N-nitro-L-arginine-methyl ester. Administration of SDF-1alpha into wounds reversed the EPC homing impairment and, with hyperoxia, synergistically enhanced EPC mobilization, homing, and wound healing. Thus, hyperoxia reversed the diabetic defect in EPC mobilization, and SDF-1alpha reversed the diabetic defect in EPC homing. The targets identified, which we believe to be novel, can significantly advance the field of diabetic wound healing.

Hyperbaric oxygen and bone marrow-derived endothelial progenitor cells in diabetic wound healing.

Endothelial progenitor cells (EPCs) are the key cellular effectors of postnatal vasculogenesis and play a central role in wound healing. In diabetes, there is a significant impairment in the number and function of circulating and wound-tissue EPC. Recent evidence indicates, that tissue-level hyperoxia achieved by therapeutic hyperbaric oxygen protocols (HBO2) can increase the mobilization of EPC from the bone marrow into peripheral blood. In this paper we review the recent reports on hyperoxia-mediated mobilization of bone marrow-derived EPC and postulate avenues of future research in this area as it applies to improving healing in chronic wounds affected by diabetes and peripheral arterial disease (PAD).

Endothelial progenitor cell release into circulation is triggered by hyperoxia-induced increases in bone marrow nitric oxide.

Endothelial progenitor cells (EPC) are known to contribute to wound healing, but the physiologic triggers for their mobilization are often insufficient to induce complete wound healing in the presence of severe ischemia. EPC trafficking is known to be regulated by hypoxic gradients and induced by vascular endothelial growth factor-mediated increases in bone marrow nitric oxide (NO). Hyperbaric oxygen (HBO) enhances wound healing, although the mechanisms for its therapeutic effects are incompletely understood. It is known that HBO increases nitric oxide levels in perivascular tissues via stimulation of nitric oxide synthase (NOS). Here we show that HBO increases bone marrow NO in vivo thereby increasing release of EPC into circulation. These effects are inhibited by pretreatment with the NOS inhibitor l-nitroarginine methyl ester (l-NAME). HBO-mediated mobilization of EPC is associated with increased lower limb spontaneous circulatory recovery after femoral ligation and enhanced closure of ischemic wounds, and these effects on limb perfusion and wound healing are also inhibited by l-NAME pretreatment. These data show that EPC mobilization into circulation is triggered by hyperoxia through induction of bone marrow NO with resulting enhancement in ischemic limb perfusion and wound healing.

The bone marrow-derived endothelial progenitor cell response is impaired in delayed wound healing from ischemia.

OBJECTIVE: Vasculogenesis relies on the recruitment of bone marrow-derived endothelial progenitor cells (BMD EPCs) and is stimulated by tissue-level ischemia. We hypothesized that the BMD EPC response is impaired in ischemic wounds and studied the relationship between BMD EPCs and wound healing. METHODS: We used transgenic Tie-2/LacZ mice, which carry the beta-galactosidase (beta-gal) reporter gene under Tie-2 promoter control. Wild-type mice were lethally irradiated and reconstituted with Tie-2/LacZ bone marrow. Four weeks later, the mice underwent unilateral femoral artery ligation/excision and bilateral wounding of the hindlimbs. Ischemia was confirmed and monitored with laser Doppler imaging. A subset of mice received incisional vs excisional nonischemic bilateral hindlimb wounds, without femoral ligation. Excisional wound closure was measured by using daily digital imaging and software-assisted calculation of surface area. RESULTS: Ischemia resulted in significantly delayed wound healing and differentially affected the number of BMD EPCs recruited to wound granulation tissue and muscle underlying the wounds. At 3 days postwounding, the granulation tissue of the wound base contained significantly fewer numbers of BMD EPCs in ischemic wounds compared with the nonischemic wounds (P < .05). In contrast, significantly more BMD EPCs were present in the muscle underlying the ischemic wounds at this same time point compared with the muscle under the nonischemic wounds (P < .05). In ischemic wounds, eventual wound closure significantly correlated with a delayed rise in BMD EPCs within the wound granulation tissue (Kendall's correlation, -.811, P = .0005) and was significantly associated with a gradual recovery of hindlimb perfusion (P < .0001). By 7 days postwounding, BMD EPCs were incorporated into the neovessels in the granulation tissue. At 14 days and 75 days, BMD EPCs were rarely observed within the wounds. CONCLUSIONS: Granulation tissue of excisional ischemic wounds showed significantly less BMD EPCs 3 days postwounding, in association with significantly delayed wound closure. However, the number of BMD EPCs were increased in ischemic hindlimb skeletal muscle, consistent with the notion that ischemia is a powerful signal for vasculogenesis. To our knowledge, this is the first report identifying a deficit in BMD EPCs in the granulation tissue of ischemic skin wounds and reporting the key role for these cells in both ischemic and nonischemic wound healing.

Stem cell mobilization by hyperbaric oxygen.

We hypothesized that exposure to hyperbaric oxygen (HBO(2)) would mobilize stem/progenitor cells from the bone marrow by a nitric oxide (*NO) -dependent mechanism. The population of CD34(+) cells in the peripheral circulation of humans doubled in response to a single exposure to 2.0 atmospheres absolute (ATA) O(2) for 2 h. Over a course of 20 treatments, circulating CD34(+) cells increased eightfold, although the overall circulating white cell count was not significantly increased. The number of colony-forming cells (CFCs) increased from 16 +/- 2 to 26 +/- 3 CFCs/100,000 monocytes plated. Elevations in CFCs were entirely due to the CD34(+) subpopulation, but increased cell growth only occurred in samples obtained immediately posttreatment. A high proportion of progeny cells express receptors for vascular endothelial growth factor-2 and for stromal-derived growth factor. In mice, HBO(2) increased circulating stem cell factor by 50%, increased the number of circulating cells expressing stem cell antigen-1 and CD34 by 3.4-fold, and doubled the number of CFCs. Bone marrow *NO concentration increased by 1,008 +/- 255 nM in association with HBO(2). Stem cell mobilization did not occur in knockout mice lacking genes for endothelial *NO synthase. Moreover, pretreatment of wild-type mice with a *NO synthase inhibitor prevented the HBO(2)-induced elevation in stem cell factor and circulating stem cells. We conclude that HBO(2) mobilizes stem/progenitor cells by stimulating *NO synthesis.

Normal human fibroblasts enable melanoma cells to induce angiogenesis in type I collagen.

BACKGROUND: We previously reported that fibroblasts induce human microvascular endothelial cells (HMVECs) to differentiate from monolayer to capillarylike morphology. We now test the hypothesis that fibroblasts modulate angiogenesis in melanoma cells. METHODS: We tested 12 human melanoma lines (2 radial growth phase (RGP), 3 vertical growth phase (VGP), and 7 metastatic (MM)) for ability to induce HMVECs to invade/migrate into collagen and form capillarylike networks. HMVEC monolayers were overlaid with 3-dimensional collagen gels embedded with melanoma cells alone (M), fibroblasts alone (F), or a 1:1 mixture of the 2 cells (M+F). After 5 days, gels were removed, fixed, and HMVEC networks were quantified by von Willebrand's factor (vWF) immunofluorescence. The influence of soluble factors on HMVEC invasion/migration into collagen was assessed with the use of acellular 3-D collagen gels overlaid on HMVEC monolayers, cultured with conditioned media (CM) derived from monolayers of M, F, or M+F. Angiogenic growth factors involved in the observed invasion/migration were identified with the use of a RayBio Cytokine Antibody Array (RayBiotech, Norcross, Ga). RESULTS: Cell line-specific variability in melanoma-supported angiogenesis was observed only when in combination with fibroblasts (analysis of variance [ANOVA], P < .01). Melanoma plus fibroblasts uniformly resulted in a significantly higher angiogenic response than melanoma alone (P < .05). One vertical growth phase and one metastatic melanoma line, while weakly angiogenic alone, induced significantly higher angiogenesis than either fibroblast or melanoma alone (P < .05) when combined with fibroblasts. CM from M or M+F induced significantly less HMVEC invasion/migration into collagen than CM from fibroblasts alone. Interleukin 8, monocyte chemotactic protein-1, and tissue inhibitor of metalloproteinase-2 were identified as significantly elevated in the media derived from M+F cultures, compared with either cell type alone. CONCLUSION: To our knowledge, this is the first report demonstrating that melanoma-supported angiogenesis in collagen is more significantly influenced by normal skin-derived fibroblasts than by the intrinsic biology of the melanoma cell type. Interleukin 8, monocyte chemotactic protein-1, and tissue inhibitor of metalloproteinase-2 are implicated as potential paracrine factors regulating this observed effect.