Harnessing tumor immunity with cytotoxics: T cells monitoring in mice bearing lung tumors treated with anti-VEGF and pemetrexed-cisplatin doublet.

BACKGROUND: Successful immunotherapy is restricted to some cancers only, and combinatorial strategies with other drugs could help to improve their efficacy. Here, we monitor T cells in NSCLC model after treatment with cytotoxics (CT) and anti-VEGF drugs, to understand when immune checkpoint inhibitors should be best associated next. METHODS: In vivo study was performed on BALB/c mice grafted with KLN205 cells. Eight treatments were tested including control, cisplatin and pemetrexed as low (LD CT) and full (MTD CT) dose as single agents, flat dose anti-VEGF and the association anti-VEGF + CT. Full immunomonitoring was performed by flow cytometry on tumor, spleen and blood over 3 weeks. RESULTS: Immunomodulatory effect was dependent upon both treatments and time. In tumors, combination groups shown numerical lower Treg cells on Day 21. In spleen, anti-VEGF and LD CT group shown higher CD8/Treg ratio on Day 7; on Day 14, higher T CD4 were observed in both combination groups. Finally, in blood, Tregs were lower and CD8/Treg ratio higher, on Day 14 in both combination groups. On Day 21, CD4 and CD8 T cells were higher in the anti-VEGF + MTD CT group. CONCLUSIONS: Anti-VEGF associated to CT triggers notable increase in CD8/Tregs ratio. Regarding the scheduling, a two-week delay after using anti-VEGF and CT could be the best sequence to optimize antitumor efficacy.

Benchmarking a Center of Excellence in Vascular Surgery: Using Acute Physiology and Chronic Health Evaluation II to Validate Outcomes in a Tertiary Care Institute.

OBJECTIVE: The Society of Vascular Surgery (SVS) has made it a top priority to implement verification of vascular "centers of excellence". Our institutional aortic network was established in 2008 in order to standardize care of patients with suspected acute aortic pathology. The implementation and success of this program has been previously reported. We sought to use our experience as a benchmark for which to develop prognostic modeling to quantify clinical status upon admission and help predict outcomes. Our objective was to validate the Acute Physiology and Chronic Health Evaluation (APACHE) II scoring system using a cohort of aortic emergencies transferred by an organized transfer network. METHOD: This was a retrospective, single institution review of patients transferred through an institutional aortic network for acute aortic pathology from 2017-2018. Demographics, comorbidities, aortic diagnosis, APACHE II score, as well as 30-day mortality were recorded. Associations with 30-day mortality were evaluated using two-sample t-tests, ANOVA models, Pearson chi-square tests and Fisher exact tests. Receiver operating characteristic (ROC) curves were fit overall and by pathology to predict 30-day mortality by Apache II total score. RESULTS: There were 395 consecutive transfers were identified. The mean age was 64.7 years. Diagnoses included Type A Dissection (n = 134), Type B (n = 81), Aortic Aneurysm (n = 122), and PAU/IMH (n = 27). Mean APACHE II score on arrival was 12. Overall there were 53 deaths (13.4%) in the cohort. Patients that died had significantly higher Apache II total scores (11.3 vs 16.5, P < .001). The area under the receiver operator characteristic (ROC) curve (AUC) was .66 for the full cohort, indicating a poor clinical prediction test. CONCLUSION: APACHE II score is a poor predictor of 30-day mortality in a large transfer network accepting all aortic emergencies. The authors believe further refining a prognostic model for diverse population will not only help in predicting outcomes but to objectively quantify illness severity in order to have a basis for comparison among institutions and verification of "centers of excellence".

Prototyping Trastuzumab Docetaxel Immunoliposomes with a New FCM-Based Method to Quantify Optimal Antibody Density on Nanoparticles.

Developing targeted nanoparticles is a rising strategy to improve drug delivery in oncology. Antibodies are the most commonly used targeting agents. However, determination of their optimal number at the surface remains a challenging issue, mainly due to the difficulties in measuring precisely surface coating levels when prototyping nanoparticles. We developed an original quantitative assay to measure the exact number of coated antibodies per nanoparticle. Using flow cytometry optimized for submicron particle analysis and beads covered with known amounts of human IgG-kappa mimicking various amounts of antibodies, this new method was tested as part of the prototyping of docetaxel liposomes coated with trastuzumab against Her2+ breast cancer. This quantification method allowed to discriminate various batches of immunoliposomes depending on their trastuzumab density on nanoparticle surface (i.e., 330 (Immunoliposome-1), 480 (Immunoliposome-2) and 690 (Immunoliposome-3), p = 0.004, One-way ANOVA). Here we showed that optimal number of grafted antibodies on nanoparticles should be finely tuned and highest density of targeting agent is not necessarily associated with highest efficacy. Overall, this new method should help to better prototype third generation nanoparticles.

Initial 200 cases of carotid artery stenting using a reversal-of-flow cerebral protection device.

AIM: Because embolic complications can occur during carotid angioplasty and stenting (CAS), a new device, the Parodi Anti-Emboli System (PAES) was developed to protect the brain from embolization. We describe our initial experience with this device. METHODS: Between September 1999 and December 2003, CAS was performed in 200 consecutive patients (146 men; mean age, 70.4 years) with symptomatic (52%) or asymptomatic (48%) severe carotid artery stenosis (>70%). De novo lesions were present in 169 patients, restenosis in 18, and radiation-induced stenosis in 13. Wallstents were inserted in all cases, with selective predilatation, and the PAES was employed during all CAS procedures. Patients were evaluated by a neurologist before and after CAS. Minor strokes, major or fatal strokes, and myocardial infarctions that occurred within 30 days of the procedure were recorded. RESULTS: The overall technical success rate for CAS using the PAES (with the PAES placed in position percutaneously) was 99%. The overall perioperative stroke and death rate was 1.5%. There were four transient neurologic events after CAS, three of which were related to hemodynamic instability and one to postoperative embolization. CONCLUSIONS: Our experience indicates that CAS using the PAES is safe and effective. The protection device may prevent the debris released by angioplasty from entering the cerebral circulation. Additional studies of this device are warranted.

Abdominal aortic aneurysm repair in high-risk and elderly patients.

Elective open repair of abdominal aortic aneurysm (AAA) is a proven surgical therapy with acceptable rates of perioperative mortality. Open AAA repair in elderly and high-risk patients, however, carries a significantly greater risk of surgical mortality and perioperative complications. Given the steady increase of life expectancy in developed nations, assessment of surgical outcomes and clarification of the role of emerging therapies in the aging population are of significant interest to the vascular surgeon. Selection of treatment options for these patients must be based on an individual approach, and assessment of outcomes must include more subtle parameters, such as quality of life, in addition to operative survival. Recent studies assessing the applicability of endoluminal graft repair in the elderly demonstrate that this avenue of treatment may offer substantial benefit to selected patients. We review the historical data regarding operative aneurysm repair in the high-risk and elderly population, and examine the impact of endoluminal therapy of AAAs in these challenging patients.

Evaluation of 5 types of fishing material, 2 sterilization methods, and a crimp-clamp system for extra-articular stabilization of the canine stifle joint.

OBJECTIVE: To evaluate the mechanical properties of 5 types of fishing material, 2 sterilization methods, and a commercially designed crimp-clamp system for the extra-articular repair of the canine stifle joint. STUDY DESIGN: Experimental study. SAMPLE POPULATION: Animals were not used in this study. METHODS: Two brands of monofilament nylon fishing line and 3 brands of monofilament nylon leader line were used to determine the effect of steam and ethylene oxide sterilization on strength and elongation of the material. A strand of 36-kg test monofilament nylon fishing material was wrapped around 2 rods or knotted to form a loop around 2 rods on a materials-testing machine. Ten trials of each brand of unsterilized, steam-sterilized, and ethylene oxide-sterilized fishing material were tested. A strand of each material was elongated to failure at a constant displacement of 1,000 mm/min to determine strength. A strand of each material was cycled 10 times to a load of 50 N to determine percent elongation. The brand of fishing material with the greatest strength and least elongation was crimped to form a loop around 2 rods on a materials-testing machine and tested as described above. ANOVA was used to determine the effect of sterilization method, brand of material, knot, wrap, and crimp on strength and elongation of the material, and a post-hoc t test was used when significant differences were found. A Student t test was used to compare fixation techniques (wrap, knot, and crimp). RESULTS: Sterilization by steam or ethylene oxide had no significant effect on the strength of the nylon fishing material. Steam sterilization resulted in significant increases (2- to 4-fold) in elongation of most nylon fishing material when compared with unsterilized material. Ethylene oxide sterilization had minimal effect on elongation of the fishing material. Mason leader line showed no significant change in strength or elongation regardless of sterilization method. Significantly less strength and significantly less elongation were demonstrated in Mason leader line that was crimped as compared with Mason leader line that was knotted. CONCLUSION: Ethlylene oxide was the preferred method of sterilization to preserve strength and minimize elongation of the fishing material. Of the materials tested, Mason leader line had the least elongation and the greatest preservation of strength when ethylene oxide was used as the sterilization method. Mason leader line and Sufix fishing line were comparable choices when steam was used as the sterilization method. Significantly less elongation was demonstrated in crimped Mason leader line as compared with knotted Mason leader line. CLINICAL RELEVANCE: Of the materials tested, Mason leader line and Sufix fishing line had the best mechanical properties for extracapsular stabilization of the canine stifle joint. Crimping is an attractive alternative to knotting and results in a reduction in elongation of the nylon fishing material.

Endoluminal graft repair for abdominal aortic aneurysms in high-risk patients and octogenarians: is it better than open repair?

OBJECTIVE: To analyze the short-term and midterm results of open and endoluminal repair of abdominal aortic aneurysms (AAA) in a large single-center series and specifically in octogenarians. METHODS: Between January 1997 and October 2000, 470 consecutive patients underwent elective repair of AAA. Conventional open repair (COR) was performed in 210 patients and endoluminal graft (ELG) repair in 260 patients. Ninety of the patients were 80 years of age or older; of these, 38 underwent COR and 52 ELG repair. RESULTS: Patient characteristics and risk factors were similar for both the entire series and the subgroup of patients 80 years or older. The overall complication rate was reduced by 70% or more in the ELG versus the COR groups. The postoperative death rate was similar for the COR and ELG groups in the entire series and lower (but not significantly) in the ELG 80 years or older subgroup versus the COR group. The 36-month rates of freedom from endoleaks, surgical conversion, and secondary intervention were 81%, 98.2%, and 88%, respectively. CONCLUSION: The short-term and midterm results of AAA repair by COR or ELG are similar. The death rate associated with this new technique is low and comparable, whereas the complication rate associated with COR in all patients and those 80 years or older in particular is greater and more serious than ELG repair. Long-term results will establish the role of ELG repair of AAA, especially in elderly and high-risk patients.

Increased expression of HDJ-2 (hsp40) in carotid artery atherosclerosis: a novel heat shock protein associated with luminal stenosis and plaque ulceration.

PURPOSE: Evidence suggests that both humoral and cellular autoimmune processes directed toward heat shock proteins (hsp) contribute to the pathogenesis of atherosclerosis. We characterized a human hsp distinct from those previously characterized in atherosclerotic lesions, termed HDJ-2. METHODS: To determine the role of HDJ-2 in atherosclerosis, we compared the level of HDJ-2 mRNA expression with the level of hsp60 and hsp70 mRNA expression in 26 carotid endarterectomy specimens and 17 normal arteries. The level of expression of HDJ-2 mRNA was also correlated to the presence of plaque ulceration and the degree of luminal stenosis associated with the lesion. RESULTS: The expression of HDJ-2 and hsp70 was significantly higher in carotid artery plaques as compared with normal arteries: HDJ-2, 6.7 +/- 1.6 vs 0.1 +/- 0.04, (P =.001); hsp70, 9.5 +/- 0.9 vs 3.7 +/- 0.8, (P =.002). There was no significant difference in hsp60 expression between carotid artery plaques and normal arteries (21.0 +/- 0.9 vs 20.6 +/- 0.8, P =.65). Increased HDJ-2 expression in carotid artery plaques was independent of hsp70 (Pearson correlation, r = 0.11; Bartlett chi(2) analysis, P =.71). Within the ulcerated plaque group, there was a correlation between degree of stenosis and high HDJ-2 mRNA expression (r = 0.896, P =.016). However, there was no correlation between degree of stenosis and high HDJ-2 mRNA expression within the nonulcerated plaque group (r = 0.530, P =.076) or within the entire group of patients (r = 0.0085, P =.97). CONCLUSION: These results demonstrate that expression of HDJ-2 is significantly increased in atherosclerotic carotid artery plaques as compared with hsp60 and hsp70 and correlates with luminal stenosis in ulcerated atherosclerotic carotid artery plaques.

Granulosa cell tumor in a guinea pig.

Abdominal distention is a common clinical sign in guinea pigs and may have many causes. Abdominal ultrasonography may be a useful diagnostic tool in differentiation of abdominal disorders in guinea pigs. Ovariohysterectomy is indicated for granulosa cell tumors and cystic rete ovarii in guinea pigs.

Evolution of vascular fellowship training in the new era of endovascular techniques.

PURPOSE: The endovascular technique has revolutionized the treatment of infrarenal abdominal aortic aneurysm (AAA). At our institution, we examined the impact of an endovascular program on the traditional operative training of the vascular fellows in the treatment of infrarenal AAA. METHODS: We examined the records of our vascular fellows' experience from July 1995 to May 2000. We introduced the endovascular treatment for infrarenal AAA in 1995. RESULTS: The fellows have performed increasing numbers of endovascular cases each year, with a predicted number of 124 cases for 1999-2000. However, despite an increase in the overall volume of patients with infrarenal AAA (102 cases in 1998-1999 and a predicted 160 cases in 1999-2000), the trainees will experience a reduction in the number of open AAAs from 61 cases in 1998-1999 to a predicted 36 cases in 1999-2000. However, the volume of open suprarenal AAA has also increased from eight cases in 1998 to 1999 to a predicted 24 cases in 1999-2000. With no significant change in the open aortoiliac occlusive cases from previous years, the current fellows will graduate with a similar volume of open aortic procedures as their predecessors. CONCLUSION: With the recent advances in endovascular technology, our traditional operative approach to the treatment of AAA disease may be lacking in the training of future vascular surgeons. At our institution, although fewer open infrarenal AAA cases were performed, the trainees have maintained the open aortic experience by performing an increased volume of suprarenal AAAs. We have to critically reevaluate and redefine what constitutes adequate vascular fellow experience in the surgical treatment of abdominal aortic aneurysms.

The Hemobahn endoprosthesis: a self-expanding polytetrafluoroethylene-covered endoprosthesis for the treatment of peripheral arterial occlusive disease after balloon angioplasty.

Endoluminal management of occlusive arterial disease has previously been limited to balloon angioplasty, either alone or with stent placement. This article discusses the Hemobahn endoprosthesis, a polytetraflouroethylene-covered nitinol stent graft. The Hemobahn device design characteristics, Food and Drug Administration phase I feasibility trial design and results, phase II pivotal trial design, and single-site phase II trial results are reviewed. The long-term outcomes of patients treated with angioplasty and Hemobahn stent grafting will determine the role of stent grafting in the management of occlusive arterial lesions below the aortic bifurcation.

Factors increasing the mortality rate for patients with ruptured abdominal aortic aneurysms.

The objective of this report was to analyze the current surgical results of operative treatment in patients suffering ruptured AAA (abdominal aortic aneurysms) and to define those independent predictive factors for mortality. During a period of 2 years, from January 1996 to December 1997, 144 patients operated on for ruptured AAA in 10 hospitals were included in a multicenter retrospective study. Among the collected variables concerning each patient, those with potential relation to surgical mortality were studied: gender, age, diabetes, hypertension, cardiopathy, pulmonary obstructive disease, preoperative renal dysfunction, symptomatic cerebrovascular disease, peripheral vascular disease, hematocrit on admission, preoperative hypotension < 80 mmHg, loss of consciousness, cardiac arrest, aortic aneurysm location (infrarenal versus non-infrarenal), iliac involvement, aneurysm size, type of rupture, left renal vein ligature, ligature of a patent inferior mesenteric artery, place of aortic cross-clamping, type of grafting, exclusion of both hypogastric arteries, venous technical complications, associated surgery, use of cell saver, intraoperative blood loss, and postoperative complications (renal failure, sepsis, coagulopathy, cardiac complications, pulmonary complications, colon ischemia, prosthetic graft complications, and need for reoperation). Those variables with statistical significance in the univariate analysis were introduced into a multivariate logistic regression model to determine the independent predictors of death. From our results we concluded that surgery for ruptured abdominal aortic aneurysms continues to have an excessively high mortality rate. Even though some preoperative variables could be identified as predictors of mortality, an absolute mortality risk has not yet been determined and the decision to negate surgery should be individualized rather than taken on that basis only. Early diagnosis and treatment of symptomatic aneurysms would improve mortality figures and selective screening should be contemplated.

Preoperative treatment with doxycycline reduces aortic wall expression and activation of matrix metalloproteinases in patients with abdominal aortic aneurysms.

PURPOSE: Matrix metalloproteinases (MMPs) are considered to play a central role in the pathogenesis of abdominal aortic aneurysms (AAAs). Doxycycline (Dox) has direct MMP-inhibiting properties in vitro, and it effectively suppresses the development of elastase-induced AAAs in rodents. The purpose of this study was to determine if treatment with Dox suppresses MMPs within human aneurysm tissue and to elucidate the molecular mechanisms underlying this effect. METHODS: Aneurysm tissues were obtained from 15 patients with an AAA, eight of whom had been treated with Dox before surgery (100 mg orally twice a day for 7 days). Protein extracts were examined by means of gelatin zymography and immunoblot analysis, and RNA was examined by means of reverse transcription-polymerase chain reaction (RT-PCR). The effects of Dox on MMP production were further examined in human THP-1 mononuclear phagocytes in vitro. RESULTS: No detectable difference was found between groups by using substrate zymography as a means of assessing total MMP activity, but Dox treatment was associated with a slight (24.4%) reduction in the activated fraction of 72-kDa gelatinase (MMP-2; P <.05). In contrast, a 2.5-fold reduction in the amount of extractable 92-kDa gelatinase (MMP-9) protein in Dox-treated patients was revealed by means of immunoblot analysis (P <.05). Also, a 5.5-fold (81.9%) reduction in MMP-9 messenger RNA (mRNA) in Dox-treated patients was demonstrated by means of quantitative competitive RT-PCR (mean +/- SE, mol MMP-9/mol beta-actin: 1.3 +/- 0.5 vs 7.2 +/- 3.1; P <.04). There was no significant difference between groups in the relative expression of MMP-2 protein or mRNA. In cultured THP-1 monocytes stimulated with phorbol ester, the expression of MMP-9 protein and mRNA were both decreased after exposure to relevant concentrations of Dox in vitro. CONCLUSION: In addition to its recognized effects as a direct MMP antagonist, Dox may influence connective tissue degradation within human aneurysm tissue by reducing monocyte/macrophage expression of MMP-9 mRNA and by suppressing the post-translational processing (activation) of proMMP-2. Through this complementary combination of mechanisms, treatment with Dox may be a particularly effective strategy for achieving MMP inhibition in patients with an AAA.

Differential expression of prostaglandin E2 and interleukin-6 in occlusive and aneurysmal aortic disease.

BACKGROUND: Both aortoiliac occlusive disease (AIOD) and abdominal aortic aneurysm disease (AAA) are traditionally considered degenerative conditions that are caused by atherosclerosis. Although it is becoming apparent that the pathophysiology of each condition has its own determinants, inflammation is thought to play a role in each. The purpose of this study was to analyze the inflammatory cytokines interleukin-6 (IL-6) and prostaglandin E2 (PGE2) in aortic disease and compare AAA with AIOD, as well as to compare both with normal aorta. METHODS: Aortic tissue was harvested at the time of aortic reconstructive surgery for AAA (n = 13) and AIOD (n = 14) or at time of organ harvest for normal (n = 16) aortic specimens. Whole organ cultures were immediately established, and the culture medium was collected after 72 hours. An enzyme-linked immunosorbent assay was used to assay for PGE2 and a lymphoproliferative assay was used to quantitate IL-6. Statistical analysis was performed using paired 2-tail t tests. RESULTS: Normal aorta expressed much less PGE2 (384 +/- 67 ng/mL) than either AAA (11,093 +/- 7,411 ng/mL) (P < .001) or AIOD (13, 719 +/- 3,355 ng/mL) (P < .002). However there was no statistically significant difference in PGE2 expression between the AAA and AIOD groups (P = . 44). The IL-6 assay also showed that normal aorta had very little expression (1,861 +/- 334 U/mL) compared with either AAA (14,329 +/- 4,159 U/mL) (P = . 02) or AIOD (39,805 +/- 8,426) (P < .001). Comparison between AAA and AIOD revealed significantly higher expression of IL-6 by the AIOD cultures (P = .03). CONCLUSIONS: AAA and AIOD are associated with increased expression of the proinflammatory cytokines PGE2 and IL-6. However, AIOD is associated with a much higher level of IL-6 expression than is AAA, although the level of PGE2 expression is the same. This differential expression of IL-6 may help explain the pathogenesis of these 2 distinct aortic diseases.

Indomethacin inhibits expansion of experimental aortic aneurysms via inhibition of the cox2 isoform of cyclooxygenase.

PURPOSE: Cyclooxygenase, either the cox1 or cox2 isoform, controls synthesis of prostaglandin E2 (PGE2), which regulates expression of matrix metalloprotease-9 (MMP-9). PGE2 and MMP-9 are elevated in aortic aneurysms. The mechanisms and time course of the inhibition of aneurysm expansion with a nonspecific cyclooxygenase inhibitor, indomethacin, were determined in an animal model. METHODS: Rats underwent aortic perfusion with saline (n = 40) as controls or with elastase. Elastase-treated animals received no treatment (n = 82) or received indomethacin (n = 73). Aortic diameters were determined at the time of aortic perfusion and when the rats were killed. The aortas were harvested and used for whole organ culture, substrate gel zymography, or histologic analysis. RESULTS: The control group demonstrated little change in aortic diameter. All the elastase-only animals developed aneurysms (maximal aortic diameter, 5.27 +/- 2.37 mm on day 14). Indomethacin markedly decreased the rate of aortic expansion (maximum aortic diameter, 3.45 +/- 1.11 mm; P <.001 vs the elastase-only group). The enzyme-linked immunosorbent assay of aortic explant culture media showed that PGE2 synthesis paralleled aortic expansion, and indomethacin decreased PGE2 synthesis. Histologically, the aortic elastin architecture was destroyed in the elastase group, but was preserved with indomethacin treatment. In situ, hybridization for cox1 and cox2 showed that cox2, but not cox1, was expressed and was co-localized by immunohistochemistry to macrophages associated with the aortic wall. Decreased levels of MMP-9 activity with indomethacin were shown by means of substrate zymography. MMP-9 was also localized to macrophages. CONCLUSION: Indomethacin attenuates aneurysm growth, and its effects are mediated via inhibition of the cox2 isoform of cyclooxygenase, which decreases PGE2 and MMP-9 synthesis.

Stress induces neurogenesis in non-neuronal cell cultures of adult olfactory epithelium.

Among the basal cells of the olfactory epithelium is a stem cell which divides and whose progeny differentiate into new sensory neurons throughout adult life. Olfactory neurogenesis is highly regulated, for example it is stimulated by epithelial damage. Previous reports implicate several growth factors in progenitor cell proliferation and neuronal differentiation in vitro but these studies differ in growth conditions and age of donors making it difficult to determine precisely the roles of neurogenic stimuli and their sites of action. The aims of the present study were to develop purified basal cell cultures from adult olfactory epithelium and to stimulate neurogenesis in defined growth conditions in order to elucidate the cellular mechanisms by which neurogenesis is stimulated after epithelial damage. We show here that differentiated olfactory sensory neurons arise after biochemical or mechanical stress of rat and mouse olfactory epithelial cell cultures in the absence of growth factors, complex media (e.g., serum, conditioned media, pituitary and hypothalamic extracts), or other cells (e.g., explants, feeder layers of glia, or other non-epithelial cells). Prior to the stress, these cultures contained basal cells and supporting cells but not neurons. After the stress, some cells differentiated into bipolar neurons expressing a number of neuronal proteins including olfactory marker protein. Bromodeoxyuridine experiments show that the differentiated neurons arose from recently divided cells which did not divide again before differentiating. We conclude that stress disrupts cell surface contacts to induce the immediate neuronal precursors to undergo final differentiation into olfactory sensory neurons. This may be a mechanism for enhanced neurogenesis after epithelial damage.

Source of elastin-degrading enzymes in mycotic aortic aneurysms: bacteria or host inflammatory response?

Elastolytic matrix metalloproteinases play a central role in the development of chronic atherosclerotic aortic aneurysms, but mycotic aortic aneurysms are a distinct and unusual form of aneurysm disease caused by bacterial infection. Mycotic aortic aneurysms follow a more rapid and unpredictable course than chronic aneurysm disease and they exhibit a predilection for the suprarenal aorta, further implying unique pathophysiologic mechanisms. The purpose of this study was to examine the nature and source of elastin-degrading enzymes in mycotic aortic aneurysm. Bacterial isolates and aortic tissues were obtained from four consecutive patients undergoing surgical repair of suprarenal mycotic aortic aneurysm. Using an in vitro 3H-labeled elastin degradation assay, elastin-degrading enzyme activity was only observed in the bacteria-conditioned medium from an isolate of Pseudomonas aeruginosa. Elastin-degrading enzyme activity in the aortic tissue homogenate of this patient was abolished by the serine protease inhibitor, phenylmethylsulfonyl fluoride, but it was not suppressed by the metalloproteinase inhibitor, ethylenediamine tetraacetic acid (EDTA). In contrast, elastin-degrading enzyme activity in the bacterial-conditioned medium was decreased by about half by both phenylmethylsulfonyl fluoride and EDTA. Elastin substrate zymography revealed two phenylmethylsulfonyl fluoride-inhibitable elastin-degrading enzyme activities in the aortic tissue homogenate that corresponded to human neutrophil elastase (approximately 30 kDa) and its stable complex with alpha 1-proteinase inhibitor (approximately 80 kDa), but no activity attributable to Pseudomonas elastase, a 33-kDa metal-dependent enzyme. Human neutrophil elastase was readily detected throughout mycotic aortic aneurysm tissues by immunohistochemistry, but elastolytic metalloproteinases were only occasionally observed. The results of this study suggest that the elastin-degrading enzyme produced in mycotic aortic aneurysm are largely serine proteases of host neutrophil origin, rather than elastases produced by the infecting microorganisms or the macrophage-derived metalloproteinases typically observed in atherosclerotic aneurysm disease. Further studies will be needed to extend these findings to a larger number of patients with mycotic aortic aneurysm and those caused by additional microorganisms.

Surgical treatment of patients with symptomatic vertebrobasilar insufficiency.

We report our experience with surgical management of symptomatic vertebrobasilar insufficiency (VBI). Forty revascularizations were carried out in 39 patients over 90 months. Dizziness (52%) and syncope/presyncope (32%) were the most common symptoms. Arteriography was performed in all patients, with subclavian steal seen in 55% of patients. Procedures performed included 22 cases of carotid-subclavian bypass or transposition (55%), seven direct vertebral reconstructions (17.5%), four great vessel reconstructions (10%), four isolated carotid endarterectomies (10%), and three axilloaxillary bypasses (7.5%). One patient died, and the combined morbidity and mortality rate was 15%. Outpatient follow-up was available on 37 of the 38 patients discharged alive. At a mean follow-up of 16.4 months, 34 patients had no VBI complaints. Three of four patients treated with CEA alone had persistent VBI complaints. We conclude that a variety of anatomic lesions can result in VBI symptoms, with subclavian steal being the most common. Procedures which directly correct the anatomic abnormality result in sustained symptom resolution with acceptable complication rates.

Generation of neurons from a nonneuronal precursor in adult olfactory epithelium in vitro.

Within the olfactory epithelium is a stem cell which can divide and differentiate to produce new sensory neurons. The identity of the neuronal stem cell is unknown but one candidate is the horizontal basal cell which lies adjacent to the basement membrane and expresses keratin. Previous attempts to generate mature sensory neurons from purified horizontal basal cells in vitro were unsuccessful. We show here for the first time that olfactory neurogenesis can be reproduced in vitro from partially-purified cultures of adult rat precursor cells cultivated in a serum-free medium. Rat olfactory epithelium was dissected from the nasal septum and separated from the underlying lamina propria, and its cells were dissociated and grown in a medium containing epidermal growth factor for 5 days. Immunochemistry showed that only supporting cells (SUS1-positive) and horizontal basal cells (keratin-positive) survived for this period. At day 6, the cells were stressed either by passaging them or by a simple mechanical stress. In each case, a morphological and immunological differentiation was observed within 24-48 hr. Newly formed bipolar cells were found to be S100-, glial fibrillary acidic protein (GFAP-), neural cell adhesion molecule (N-CAM+), and/or microtubule-associated protein 5 (MAP-5+). After passaging 14% of the surviving cells were immature neurons (MAP-5+) and 4% were mature olfactory neurons (MAP-5+) and olfactory marker protein (OMP+)). In addition the same experiment was conducted on transgenic mice in which the lacZ gene was linked to the OMP promoter. Using 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-Gal) staining we showed that OMP+ cells disappeared before day 5 in culture but reappeared after passaging. These results suggest that olfactory sensory neurons can arise from a non-neuronal precursor, probably the keratin-positive horizontal basal cell.