Joseph J. Cullen

Ascorbic acid: Chemistry, biology and the treatment of cancer

Since the discovery of vitamin C, the number of its known biological functions is continually expanding. Both the names ascorbic acid and vitamin C reflect its antiscorbutic properties due to its role in the synthesis of collagen in connective tissues. Ascorbate acts as an electron-donor keeping iron in the ferrous state thereby maintaining the full activity of collagen hydroxylases; parallel reactions with a variety of dioxygenases affect the expression of a wide array of genes, for example via the HIF system, as well as via the epigenetic landscape of cells and tissues. In fact, all known physiological and biochemical functions of ascorbate are due to its action as an electron donor. The ability to donate one or two electrons makes AscH(-) an excellent reducing agent and antioxidant. Ascorbate readily undergoes pH-dependent autoxidation producing hydrogen peroxide (H(2)O(2)). In the presence of catalytic metals this oxidation is accelerated. In this review, we show that the chemical and biochemical nature of ascorbate contribute to its antioxidant as well as its prooxidant properties. Recent pharmacokinetic data indicate that intravenous (i.v.) administration of ascorbate bypasses the tight control of the gut producing highly elevated plasma levels; ascorbate at very high levels can act as prodrug to deliver a significant flux of H(2)O(2) to tumors. This new knowledge has rekindled interest and spurred new research into the clinical potential of pharmacological ascorbate. Knowledge and understanding of the mechanisms of action of pharmacological ascorbate bring a rationale to its use to treat disease especially the use of i.v. delivery of pharmacological ascorbate as an adjuvant in the treatment of cancer.

Measurement of superoxide dismutase, catalase and glutathione peroxidase in cultured cells and tissue

Cells contain a large number of antioxidants to prevent or repair the damage caused by reactive oxygen species, as well as to regulate redox-sensitive signaling pathways. General protocols are described to measure the antioxidant enzyme activity of superoxide dismutase (SOD), catalase and glutathione peroxidase. The SODs convert superoxide radical into hydrogen peroxide and molecular oxygen, whereas the catalase and peroxidases convert hydrogen peroxide into water. In this way, two toxic species, superoxide radical and hydrogen peroxide, are converted to the harmless product water. Western blots, activity gels and activity assays are various methods used to determine protein and activity in both cells and tissue depending on the amount of protein required for each assay. Other techniques including immunohistochemistry and immunogold can further evaluate the levels of the various antioxidant enzymes in tissues and cells. In general, these assays require 24–48 h to complete.

Pancreatic anastomotic leak after pancreaticoduodenectomy : incidence, significance, and management

Anastomotic leak at the pancreaticojejunostomy remains a common and dreaded complication after pancreaticoduodenectomy. Our aim was to determine the incidence, presentation, methods of management, and preoperative and postoperative factors that influence the clinical outcome. We reviewed our collective experience with 375 consecutive patients undergoing pancreaticoduodenectomy from 1980 to 1992 for a variety of pathologic indications. Clinical, biochemical, intraoperative, and postoperative factors were reviewed in an attempt to determine prognostic factors. Sixty-six patients (18%) developed a pancreatic anastomotic leak as determined by increased amylase in drainage fluid (44%), radiographic documentation (41%), operative re-exploration (9%), or percutaneous drainage of a peripancreatic, amylase-containing fluid collection (6%). Most leaks (73%) were clinically insignificant and were managed by simple maintenance of intraoperatively placed drains. Active intervention was required in 18 patients (27%) and included percutaneous drainage in 8, completion pancreatectomy in 7, and reoperative drainage with or without anastomotic repair in 3. Although octreotide was used therapeutically in 13 patients (20%), a significant, objective response was noted in only 1 patient. Five (8%) of the 66 patients died, all related directly to the pancreatic leak. The overall operative mortality was lower, 15 (4%) of 375 patients. Of the clinical, biochemical, intraoperative, and postoperative factors reviewed to determine prognostic factors, only postoperative intra-abdominal hemorrhage predisposed the patient to mortality as a result of the pancreatic anastomotic leak. We conclude that most anastomotic leaks at the pancreaticojejunostomy after pancreaticoduodenectomy can be managed conservatively. Use of octreotide to aid in closure of the pancreatic leak was disappointing, whereas patients with postoperative intra-abdominal hemorrhage or those requiring completion pancreatectomy to manage the anastomotic leak have increased mortality.

Surgical management of Meckel's diverticulum: An epidemiologic, population-based study

ObjectiveThe authors determined whether Meckels diverticulum, discovered incidentally at operation, should be removed. Summary Background DataIt is not clear from the medical literature whether the risk of an incidental Meckels diverticulectomy is greater than the risk of leaving the diverticulum in place. MethodsThe authors used the medical experience of Olmsted County, Minnesota residents for the period 1950 to 1992 to answer the question. ResultsDuring the period, 58 residents developed Meckels complications that required diverticulectomies. The incidence of complications was 87 per 100.000 person-years, and the lifetime risk (to 80 years of age) of developing them was 6.4%. The risks were similar throughout the period and at all ages of life, but were greater among men (124 per 100,000 person-years) than women (50 per 100,000 person-years, p < 0.05). Diverticulectomies for complications carried an operative mortality and morbidity of 2% and 12% and a cumulative risk of long-term postoperative complications of 7%, whereas incidental diverticulectomies done in 87 residents during the period carried corresponding rates of only 1%, 2%, and 2%, respectively. ConclusionsMeckels diverticula discovered incidentally at operation should be removed for most patients, regardless of age.

Mechanisms of Ascorbate-Induced Cytotoxicity in Pancreatic Cancer

Purpose: Pharmacologic concentrations of ascorbate may be effective in cancer therapeutics. We hypothesized that ascorbate concentrations achievable with i.v. dosing would be cytotoxic in pancreatic cancer for which the 5-year survival is <3%. Experimental Design: Pancreatic cancer cell lines were treated with ascorbate (0, 5, or 10 mmol/L) for 1 hour, then viability and clonogenic survival were determined. Pancreatic tumor cells were delivered s.c. into the flank region of nude mice and allowed to grow at which time they were randomized to receive either ascorbate (4 g/kg) or osmotically equivalent saline (1 mol/L) i.p. for 2 weeks. Results: There was a time- and dose-dependent increase in measured H2O2 production with increased concentrations of ascorbate. Ascorbate decreased viability in all pancreatic cancer cell lines but had no effect on an immortalized pancreatic ductal epithelial cell line. Ascorbate decreased clonogenic survival of the pancreatic cancer cell lines, which was reversed by treatment of cells with scavengers of H2O2. Treatment with ascorbate induced a caspase-independent cell death that was associated with autophagy. In vivo, treatment with ascorbate inhibited tumor growth and prolonged survival. Conclusions: These results show that pharmacologic doses of ascorbate, easily achievable in humans, may have potential for therapy in pancreatic cancer. Clin Cancer Res; 16(2); 509–20

A Decade of Change in Obesity Surgery

Background: The International (formerly National) Bariatric Surgery Registry began collecting data in January 1986. The aim of this study was to examine changes in the practice of surgical treatment of severe obesity that occurred during the decade of 1986 through 1995, as observed in the IBSR data. Methods: All data submitted to the IBSR during the decade were transferred to the IBM mainframe computer for analysis. Characteristics of operative type populations were compared over time using analysis of variance (ANOVA) for age, body mass index (BMI), operative weight and Chi-square (χ2) test for gender. Results: There has been a steady increase over the decade in mean patient weight. The operations used have changed from predominantly ‘simple’ operations to more frequent use of ‘complex’ operations. Within the categories of ‘simple’ and ‘complex’, an increase in the variety of operations occurred. As a group, patients with ‘simple’ operations have been heavier, more often male and public pay patients than those who have undergone ‘complex’ operations. One year weight loss was greater for Roux-en-Y gastric bypass (RGB) than vertical banded gastroplasty (VBG), but follow-up rates were too low to study the relative merits of the operations used. The reported incidence of operative mortality and serious complications (leak with peritonitis, abscess and pulmonary embolism) remained low. Conclusions: These observations and their implications can be summarized in three statements which relate to action for improved patient care in the beginning of the new century: (1) increasing weight of candidates for surgical treatment during this decade indicates the need for earlier use of operative treatment before irreversible complications of obesity can develop; (2) low risk of obesity surgery, decreasing postoperative hospital stay, and early weight control support the continued and increased use of surgical treatment; (3) continued widespread use of both ‘simple’ and ‘complex’ operations with increased modifications of standard RGB and VBG procedures emphasizes the need for standardized long-term data and analyses regarding both weight control and postoperative side-effects.

A Prospective Study of Outcomes, Healthcare Resource Utilization, and Costs Associated With Postoperative Nosocomial Infections

OBJECTIVE We evaluated 4 important outcomes associated with postoperative nosocomial infection: costs, mortality, excess length of stay, and utilization of healthcare resources. DESIGN The outcomes for patients who underwent general, cardiothoracic, and neurosurgical operations were recorded during a previous clinical trial. Multivariable analyses including significant covariates were conducted to determine whether nosocomial infection significantly affected the outcomes. SETTING A large tertiary care medical center and an affiliated Veterans Affairs Medical Center. PATIENTS A total of 3,864 surgical patients. RESULTS The overall nosocomial infection rate was 11.3%. Important covariates included age, Karnofsky score, McCabe and Jackson classification of the severity of underlying disease, National Nosocomial Infection Surveillance system risk index, and number of comorbidities. After accounting for covariates, nosocomial infection was associated with increased postoperative length of stay, increased costs, increased hospital readmission rate, and increased use of antimicrobial agents in the outpatient setting. Nosocomial infection was not associated independently with a significantly increased risk of death in this surgical population. CONCLUSION Postoperative nosocomial infection was associated with increased costs of care and with increased utilization of medical resources. To accurately assess the effects of nosocomial infections, one must take into account important covariates. Surgeons seeking to decrease the cost of care and resource utilization must identify ways to decrease the rate of postoperative nosocomial infection.

Expression of antioxidant enzymes in diseases of the human pancreas: Another link between chronic pancreatitis and pancreatic cancer

Introduction Chronic pancreatitis is a significant risk factor for pancreatic cancer and is associated with the generation of reactive oxygen species. Cells contain a large number of antioxidants to prevent or repair the damage caused by reactive oxygen species. There are three major types of primary intracellular antioxidant enzymes in mammalian cells: superoxide dismutase (SOD), catalase, and peroxidase, of which glutathione peroxidase is the most prominent. Aim To determine the level of antioxidant enzymes in human pancreas from normal, chronic pancreatitis, and pancreatic cancer specimens. Methodology Immunohistochemical analysis for manganese SOD, copper/zinc SOD, catalase, and glutathione peroxidase expression using the avidin–biotin–peroxidase complex method was performed on pancreatic specimens previously fixed in formalin and embedded in paraffin. A quantitative digital imaging methodology was used to examine antioxidant staining in the pancreatic tissue. Cytoplasmic regions of ductal and acinar cells were identified and digitized. Mean gray-level pixel values were then obtained for each of these regions. Results Cytoplasmic values of manganese SOD, catalase, and glutathione peroxidase were decreased in pancreatic cells from chronic pancreatitis specimens when compared with normal pancreas. In pancreatic carcinoma specimens, mean cytoplasmic gray-level values of all antioxidant enzymes were decreased when compared with normal pancreas. Conclusion There appears to be a gradual decrease in antioxidant enzyme expression in pancreatic cells from normal pancreas to chronic pancreatitis to pancreatic cancer.

Efficacy of beta-lapachone in pancreatic cancer treatment: exploiting the novel, therapeutic target NQO1.

NAD(P)H:quinone oxidoreductase (NQO1) is elevated in human pancreatic cancers. We hypothesized that £]-lapachone, a novel 1,2-naphthoquinone with potential antitumor activity in cancer cells expressing elevated levels of NQO1, would induce cytotoxicity in pancreatic cancer cells, wherein this two-electron reductase was recently found elevated. ?-lapachone decreased clonogenic cell survival, metabolic cell viability, and anchorage-independent growth in soft agar. The cytotoxic in vitro effects of ?- lapachone were inhibited with co-administration of dicumarol, a specific inhibitor of NQO1. In pre-established human pancreatic tumor xenografts in nude mice, ?-lapachone demonstrated greater tumor growth inhibition when given intratumorally compared to when complexed with cyclodextrin to increase its bioavailability. Due to the poor prognosis of patients with pancreatic cancer and the limited effectiveness of surgery, chemotherapy, and radiation therapy, treatment regimens based on sound, tumor-specific rationales are desperately need for this disease.

2-deoxy-D-glucose causes cytotoxicity, oxidative stress, and radiosensitization in pancreatic cancer.

Glucose metabolism as assessed by (18)FDG PET imaging provides prognostic information in patients with pancreatic cancer but the implications of manipulating glucose metabolism for therapeutic purposes are unknown. Based on previous results with other cancer cell types, we hypothesized that inhibition of glucose metabolism in pancreatic cancer cells would cause cell killing via oxidative stress resulting from disruptions in thiol metabolism. 2-Deoxy-D-glucose (2DG), a chemical inhibitor of glucose metabolism, and glucose deprivation induced cytotoxicity in human pancreatic cancer cells in a time-and dose-dependent manner as well as causing significant increases in metabolic oxidative stress as measured by increased glutathione disulfide accumulation and NADP(+)/NADPH ratios. Simultaneous administration of the thiol antioxidant N-acetylcysteine protected pancreatic cancer cells against the c-ytotoxic effects of 2DG as well as reversing 2DG-induced glutathione disulfide accumulation and augmenting intracellular cysteine pools. In nude mice with heterotopic pancreatic tumors, the combination of 2DG and ionizing radiation resulted in greater inhibition of tumor growth and increased survival, relative to either agent alone. These results support the hypothesis that inhibiting glucose metabolism causes cytotoxicity in human pancreatic cancer cells via metabolic oxidative stress and disruptions in thiol metabolism. These results also support the speculation that inhibitors of glucose metabolism can be used in combination with classical oxidative stress-inducing agents (such as ionizing radiation) to enhance therapeutic responses in pancreatic cancer.