Gerald C. O’Sullivan

A prospective study of common bile duct calculi in patients undergoing laparoscopic cholecystectomy: Natural history of choledocholithiasis revisited

Objective:To define the incidence of problematic common bile duct calculi in patients undergoing laparoscopic cholecystectomy. Summary Background Data:In patients selected for laparoscopic cholecystectomy, the true incidence of potentially problematic common bile duct calculi and their natural history has not been determined. We evaluated the incidence and early natural history of common bile duct calculi in all patients undergoing laparoscopic cholecystectomy with intraoperative and delayed postoperative cholangiography. Methods:Operative cholangiography was attempted in all patients. In those patients in whom a filling defect was noted in the bile duct, the fine bore cholangiogram catheter was left securely clipped in the cystic duct for repeated cholangiography at 48 hours and at approximately 6 weeks postoperatively. Results:Operative cholangiography was attempted in 997 consecutive patients and was accomplished in 962 patients (96%). Forty-six patients (4.6%) had at least one filling defect. Twelve of these had a normal cholangiogram at 48 hours (26% possible false-positive operative cholangiogram) and a further 12 at 6 weeks (26% spontaneous passage of calculi). Spontaneous passage was not determined by either the number or size of calculi or by the diameter of the bile duct. Only 22 patients (2.2% of total population) had persistent common bile duct calculi at 6 weeks after laparoscopic cholecystectomy and retrieved by endoscopic retrograde cholangiopancreatography. Conclusions:Choledocholithiasis occurs in 3.4% of patients undergoing laparoscopic cholecystectomy but more than one third of these pass the calculi spontaneously within 6 weeks of operation and may be spared endoscopic retrograde cholangiopancreatography. Treatment decisions based on assessment by operative cholangiography alone would result in unnecessary interventions in 50% of patients who had either false positive studies or subsequently passed the calculi. These data support a short-term expectant approach in the management of clinically silent choledocholithiasis in patients selected for LC.

The Fas counterattack: cancer as a site of immune privilege

Abstract Resistance to apoptosis through the Fas receptor pathway coupled with expression of the Fas ligand might enable many cancers to deliver a pre-emptive strike or counterattack against the immune system. Therapeutic exploitation of this has exciting potential, but now seems more complex and hazardous than was first evident.

PCR detection of Mycobacterium paratuberculosis in Crohn’s disease granulomas isolated by laser capture microdissection

Background and aims: The uncertainty surrounding the role of Mycobacterium avium subsp paratuberculosis (Map) in Crohn’s disease has been compounded by possible contamination from Map present in the lumen microflora. This study used laser capture microdissection (LCM) and polymerase chain reaction (PCR) to detect Map DNA in subepithelial granulomas, isolated from 15 surgically resected, formalin fixed specimens of granulomatous Crohn’s disease and from 12 granulomatous disease controls (10 bowel, 2 non-bowel). Methods: The effect of amplicon size on reliability of PCR from formalin fixed samples was examined by amplifying 435 bp and 133 bp sequences of the human APC gene. After this, nested primers were designed to detect a small fragment (155 bp) of the Map specific IS900 gene in Crohn’s granulomas. LCM isolated granulomas from Map culture positive bovine intestine was used as positive control. PCR product specificity was confirmed by direct DNA sequencing. Results: The smaller, but not the larger, fragment of the APC gene amplified reliably in all samples. Amplification of the 155 bp fragment of the IS900 gene detected Map DNA in microdissected Crohn’s granulomas in 6 of 15 cases, and in 0 of 12 disease control granulomas. Conclusions: LCM can be used to detect Map DNA in granulomas in a proportion of patients with Crohn’s disease. However, formalin fixation requires that comparatively short DNA fragments of the Map specific IS900 gene be targeted, to permit consistent detection. Detection of Map DNA within granulomas might suggest an infectious aetiology in a subset of patients; alternatively, a transmissible agent may not be involved but mycobacterial DNA may influence pathogenesis by modifying the local cytokine responses.

Rapid responses to aldosterone in human distal colon

Aldosterone at normal physiological levels induces rapid increases in intracellular calcium and pH in human distal colon. The end target of these rapid signaling responses are basolateral K+ channels. Using spectrofluorescence microscopy and Ussing chamber techniques, we have shown that aldosterone activates basolateral Na/H exchange via a protein kinase C and calcium-dependent signaling pathway. The resultant intracellular alkalinization up-regulates an adenosine triphosphate (ATP)-dependent K+ channel (K(ATP)) and inhibits a Ca2+ -dependent K+ channel (K(Ca)). In Ussing chamber experiments, we have shown that the K(ATP) channel is required to drive sodium absorption, whereas the K(Ca) channel is necessary for both cyclic adenosine monophosphate and calcium-dependent chloride secretion. The rapid effects of aldosterone on intracellular calcium, pH, protein kinase C and K(ATP), K(Ca) channels are insensitive to cycloheximide, actinomycin D, and spironalactone, indicating a nongenomic mechanism of action. We propose that the physiological role for the rapid nongenomic effect of aldosterone is to prime pluripotential epithelia for absorption by simultaneously up-regulating K(ATP) channels to drive absorption through surface cells and down-regulating the secretory capacity by inhibiting K(Ca) channels involved in secretion through crypt cells.

Rapid effects of corticosteroids on cytosolic protein kinase C and intracellular calcium concentration in human distal colon

Recent studies from our laboratory have reported rapid (< 1 min) non-genomic activation of potassium recycling, Na+-H+ exchange, protein kinase C (PKC) activity and PKC-sensitive Ca2+ entry by mineralocorticoids in mammalian distal colonic epithelium. Previous studies from other laboratories have described stimulation of the Na+-H+ exchanger by PKC activation. Here a rapid non-genomic effect of aldosterone on PKC activity and intracellular free calcium [Ca2+]i is demonstrated in human distal colonic epithelium. Rapid activation (after 15 min incubation) of basal PKC activity was observed in cytosolic fractions of human colonic epithelium by aldosterone, fludrocortisone and deoxycorticosterone acetate (DOCA). PKC activation was inhibited by the specific PKC inhibitor bisindolylmaleimide (GF109203X). The glucocorticoid hydrocortisone failed to activate PKC activity. Aldosterone induced a rapid increase in [Ca2+]i in isolated human colonic crypts. This stimulatory effect on [Ca2+]i was inhibited by the PKC inhibitor chelerythrine chloride. Hydrocortisone and dexamethasone similarly failed to increase [Ca2+]i. These results indicate that intracellular signalling for aldosterone involves changes in [Ca2+]i via activation of PKC. Since stimulation of PKC activity and increase in [Ca2+]i are apparent at normal circulating levels of aldosterone, our findings may have important physiological implications and prompt a reassessment of mineralocorticoid effects on electrolyte homeostasis.

The emerging role of viruses in the treatment of solid tumours

There is increasing optimism for the use of non-pathogenic viruses in the treatment of many cancers. Initial interest in oncolytic virotherapy was based on the observation of an occasional clinical resolution of a lymphoma after a systemic viral infection. In many cancers, by comparison with normal tissues, the competency of the cellular anti-viral mechanism is impaired, thus creating an exploitable difference between the tumour and normal cells, as an unimpeded viral proliferation in cancer cells is eventually cytocidal. In addition to their oncolytic capability, these particular viruses may be engineered to facilitate gene delivery to tumour cells to produce therapeutic effects such as cytokine secretion and anti -tumour immune responses prior to the eventual cytolysis. There is now promising clinical experience with these viral strategies, particularly as part of multimodal studies, and already several clinical trials are in progress. The limitations of standard cancer chemotherapies, including their lack of specificity with consequent collateral toxicity and the development of cross-resistance, do not appear to apply to viral-based therapies. Furthermore, virotherapy frequently restores chemoradiosensitivity to resistant tumours and has also demonstrated efficacy against cancers that historically have a dismal prognosis. While there is cause for optimism, through continued improvements in the efficiency and safety of systemic delivery, through the emergence of alternative viral agents and through favourable clinical experiences, clinical trials as part of multimodal protocols will be necessary to define clinical utility. Significant progress has been made and this is now a major research area with an increasing annual bibliography.

Mesenteric cysts — A series of six cases with a review of the literature

BackgroundSymptomatic mesenteric cysts account for only 1 in 100,000 acute adult and 1 in 20,000 acute paediatric admissions. Acute symptoms are related to compression of intra-abdominal organs or stretching of the mesentery by rapid expansion. An abdominal mass, mobile in transverse but not longitudinal plane, is often the only physical finding.MethodWe outline the presentation, management and histological findings of 6 cases that presented to this hospital from 1987–1997.ResultsThere were 5 adults aged 32–79 yr and an 8 yr old boy. The child presented acutely with a painful tender abdominal mass. Of the adults, 1 presented acutely, 2 with chronic symptoms and 2 were incidental findings. Mesenteric cysts were successfully resected in all cases.ConclusionSurgical intervention is recommended and resection of adjacent bowel may be necessary for complete excision. Successful minimal access surgery via the laparoscope has been reported and may become more widely applicable.

Non-viral in vivo immune gene therapy of cancer: combined strategies for treatment of systemic disease

Many patients with various types of cancers have already by the time of presentation, micrometastases in their tissues and are left after treatment in a minimal residual disease state [Am J Gastroenterol 95(12), 2000]. To prevent tumour recurrence these patients require a systemic based therapy, but current modalities are limited by toxicity or lack of efficacy. We have previously reported that immune reactivity to the primary tumour is an important regulator of micrometastases and determinant of prognosis. This suggests that recruitment of specific anti-tumour mechanisms within the primary tumour could be used advantageously for tumour control as either primary or neo-adjuvant treatments. Recently, we have focused on methods of stimulating immune eradication of solid tumours and minimal residual disease using gene therapy approaches. Gene therapy is now a realistic prospect and a number of delivery approaches have been explored, including the use of viral and non-viral vectors. Non-viral vectors have received significant attention since, in spite of their relative delivery inefficiency, they may be safer and have greater potential for delivery of larger genetic units. By in vivo electroporation of the primary tumour with plasmid expressing GM-CSF and B7-1, we aim to stimulate immune eradication of the treated tumour and associated metastases. In this symposium report, we describe an effective gene based approach for cancer immunotherapy by inducing cytokine and immune co-stimulatory molecule expression by the growing cells of the primary tumour using a plasmid electroporation gene delivery strategy. We discuss the potential for enhancement of this therapy by its application as a neoadjuvant to surgical excision and by its use in combination with suppressor T cell depletion.

Lithium Modulates Autophagy in Esophageal and Colorectal Cancer Cells and Enhances the Efficacy of Therapeutic Agents In Vitro and In Vivo

Many epithelial cancers, particularly gastrointestinal tract cancers, remain poor prognosis diseases, due to resistance to cytotoxic therapy and local or metastatic recurrence. We have previously shown that apoptosis incompetent esophageal cancer cells induce autophagy in response to chemotherapeutic agents and this can facilitate their recovery. However, known pharmacological inhibitors of autophagy could not enhance cytotoxicity. In this study, we have examined two well known, clinically approved autophagy inducers, rapamycin and lithium, for their effects on chemosensitivity in apoptosis incompetent cancer cells. Both lithium and rapamycin were shown to induce autophagosomes in esophageal and colorectal cancer cells by western blot analysis of LC3 isoforms, morphology and FACS quantitation of Cyto-ID or mCherry-GFP-LC3. Analysis of autophagic flux indicates inefficient autophagosome processing in lithium treated cells, whereas rapamycin treated cells showed efficient flux. Viability and recovery was assessed by clonogenic assays. When combined with the chemotherapeutic agent 5-fluorouracil, rapamycin was protective. In contrast, lithium showed strong enhancement of non-apoptotic cell death. The combination of lithium with 5-fluorouracil or oxaliplatin was then tested in the syngenic mouse (balb/c) colorectal cancer model—CT26. When either chemotherapeutic agent was combined with lithium a significant reduction in tumor volume was achieved. In addition, survival was dramatically increased in the combination group (p < 0.0001), with > 50% of animals achieving long term cure without re-occurrence (> 1 year tumor free). Thus, combination treatment with lithium can substantially improve the efficacy of chemotherapeutic agents in apoptosis deficient cancer cells. Induction of compromised autophagy may contribute to this cytotoxicity.

Immune gene therapy as a neoadjuvant to surgical excision to control metastatic cancers

We investigated if the range of efficacy of a gene-based immunotherapy of solid tumours against systemic disease could be extended when used as a neoadjuvant to surgery. Hundred percent mice whose subcutaneous tumours were surgically removed 4 days post-electroporation with GM-CSF and B7-1 plasmid were systemically resistance to tumour rechallenge. In mice bearing both subcutaneous and hepatic tumours, neoadjuvant treatment of the primary tumour resulted in significant reduction in, or elimination of, hepatic tumour growth, with a significant increase in survival, indicating that control of the primary tumour by immunotherapy was not a prerequisite for containment of systemic disease.