[Retrieval of a migrated plastic stent in a 51-year-old man]. / Bergung eines migrierten Doppel-Pigtail-Stents bei einem 51-jährigen Patienten.

BACKGROUND: Endosonographically guided transgastric drainage is the first-line interventional therapy of walled-off necrosis and symptomatic pancreatic pseudocysts in necrotizing pancreatitis. Plastic stents or lumen apposing metal stents are commonly used. A possible complication of endoscopic therapy is stent migration. CASE REPORT: We report upon a 51-year-old man who presented with acute necrotizing pancreatitis. Transgastric necrosectomy was performed and 5 transmural double-pigtail stents (DPS) were left in situ to drain the residual retroperitoneal cavity. The patient recovered and 4 stents were endoscopically removed 5 weeks later on an outpatient basis, whereas the fifth stent was suspected to have passed spontaneously via the natural route. The asymptomatic patient presented 3 months later for follow-up computed tomography. The necrosis had healed but one DPS was seen beyond the gastric wall near the kidney. Transmural access to the stent could be achieved by an endosonographically guided puncture toward the proximal portion of the stent followed by placement of a hydrophilic guidewire alongside the stent. A new gastrostomy was created by using a 6F cystotome followed by wire-guided dilation with a 12 mm balloon. The stent could then be grasped with transmurally inserted rat-tooth forceps and repositioned across the gastrostomy site. The patient was given prophylactic antibiotics. After removal of the stent, the patient could be discharged. CONCLUSION: Herein, we present the successful endosonographically guided transmural removal of a retroperitoneally migrated plastic stent. Of note, in our patient we had to rely completely on endosonography and radiography for localization and targeting of the stent, since the former necrotic cavity had meanwhile completely healed.

Direct gene transfer into the colon using a double-balloon catheter.

BACKGROUND AND STUDY AIMS: In spite of the many advances that have been made in understanding the molecular basis for diseases, a major obstacle to the treatment of human disorders remains the inability to express genes at specific sites in vivo. Recent progress in gene transfer technology has provided access to a variety of recombinant gene products that can be applied in clinical medicine for therapeutic purposes. MATERIALS AND METHODS: In an animal model, we describe here the way in which a marker gene can be introduced into the colon using a double-balloon catheter. Cationic liposomes were used as vehicles to introduce DNA into the living organism. RSV-LacZ plasmid coding for the enzyme beta-galactosidase was used as a marker gene. Cells expressing beta-galactosidase can be stained using the chromogen X-gal. Positive cells show a blue coloration in the cytoplasm. RESULTS: Both absorptive cells and goblet cells were successfully transduced with the marker gene. No evidence of similar staining was observed in control animals receiving a control plasmid or liposomes alone. CONCLUSIONS: The method used is a simple, safe, and nontoxic way of delivering genes of interest to specific sites in the colon. Gene transfer may offer fresh potential for endoscopic interventions in colonic disease.

Colon epithelium can be transiently transfected with liposomes, calcium phosphate precipitation and DEAE dextran in vivo.

BACKGROUND: The introduction of recombinant DNA into cells is the initial step toward the development of gene therapy. It has been shown that cationic liposomes are useful vehicles to introduce DNA into colon epithelial cells in vivo. METHODS: In the present study we compared the efficacy of different nonviral transfection methods into the colon wall. In anesthetized rats, a double balloon catheter was advanced into the colon and a chloramphenicol acetyltransferase (CAT) reporter plasmid complexed to liposomes, mixed with DEAE dextran, or precipitated with calcium phosphate was instilled. Following 2 days CAT activity was determined in the transfected colon segments. RESULTS: DEAE dextran and liposomes were more effective than calcium phosphate, whereas naked DNA was not taken up by the colon epithelial cells. Reporter gene expression was dose-dependent. Expressing cell types did not differ utilizing the various transfection methods as judged by X-gal staining of colon sections after transfection with a LacZ reporter plasmid. CONCLUSION: These data indicate that in addition to liposomes, plasmid DNA mixed with DEAE dextran can be taken up by colon epithelial cells. This transfection techniques may prove useful in the development of gene therapy approaches for colon disease.

Helicobacter pylori in the oral cavity: high prevalence and great DNA diversity.

To test the hypothesis that Helicobacter pylori may be transmitted by the oral-oral route, we applied nested PCR and DNA sequencing to detect and analyze H. pylori DNA in the oral cavity of 20 adult patients undergoing endoscopy. Dental plaques of molars, premolars, and incisors and saliva were collected. Additional paraffin-embedded gastric biopsies were analyzed in four patients. Two sets of highly sensitive and specific primers, EHC-U/EHC-L and ET5-U/ET-5L directed to a 860-bp fragment of H. pylori DNA, were used in the nested PCR. Eight patients had an active infection in the stomach determined with the [13C]urea breath test and the other 12 were negative. Nested PCR showed that all 20 subjects (100%) were positive for H. pylori in the oral cavity. DNA sequencing demonstrated that all tested PCR products of the expected size from the oral samples have more than 97% identity with that from H. pylori type strain ATCC 43629. However, sequences differed in oral samples from different subjects as well as between different oral locations and gastric biopsies within the same individuals. In conclusion, the oral cavity may be a permanent reservoir for H. pylori and can harbor multiple H. pylori strains at the same time.

Liposome mediated gene transfer into the rat oesophagus.

BACKGROUND: Cancer of the oesophagus has so far eluded every attempt at pharmacological treatment. The recent advent of somatic gene therapy offers a new therapeutic approach to malignant tumours. AIM: To investigate whether and how gene transfer into the oesophagus can be achieved. METHODS: A LacZ reporter gene was used as marker and transferred into the oesophagus of rats using cationic liposomes. Gene transfer was achieved by either luminal instillation into a closed segment using a double balloon catheter, or by intramural injection through a needle. Expression of beta-galactosidase was monitored in the oesophagus and various control tissues by histochemistry, polymerase chain reaction (PCR), reverse transcriptase PCR, and Southern blotting. RESULTS: Up to 100 days after in vivo gene transfer beta-galactosidase activity could be demonstrated in the oesophagus. Following luminal application, the transgene was expressed in epithelial cells whereas intramural injection induced preferential expression in fibroblasts. CONCLUSION: In vivo gene transfer into the esophagus is feasible and safe, and the route of administration largely determines cell type specificity. This novel approach will enable in vivo studies of growth, differentiation, and malignant transformation in the oesophagus, and may open new avenues to the confinement of oesophageal malignancies.

Helicobacter pylori in dental plaque: a comparison of different PCR primer sets.

This study was designed to compare different primer sets for PCR analysis of H. pylori in the same series of 40 dental plaque samples. Three pairs of primers, HPU1/HPU2, HP1/HP2, and EHC-U/EHC-L, directed to the urease A gene, 16S rRNA gene, or 860-bp DNA of H. pylori, respectively, were used. Our results demonstrate that EHC-L/EHC-U were more specific and sensitive for H. pylori added to saliva or dental plaque than HPU1/HPU2 and HP1/HP2. The detection rates for H. pylori DNA in dental plaque samples from randomly selected adult patients from the Dental Clinic of the University of Ulm were 26.5% (9/34) for HPU1/HPU2, 78.9% (30/38) for HP1/HP2, and 100% (40/40) for EHC-U/EHC-L (P < 0.001). Nested PCR using primers directed to the 860-bp DNA of H. pylori further confirmed the presence of H. pylori DNA (40/40) in all these samples. Our results indicate that primers EHC-U/EHC-L are to be recommended for PCR detection of H. pylori in the oral cavity.

Direct gene transfer into the rat pancreas using DNA-liposomes.

BACKGROUND: Pancreatic cancer represents a malignancy with very poor clinical prognosis and limited therapeutic potential. Recent developments of gene transfer technology offer new therapeutic avenues by delivering recombinant genes directly into normal or neoplastic tissue in vivo. METHODS: Here we show that the LacZ marker gene, complexed to cationic liposomes, can be introduced into the pancreas by either intraductal or intra-arterial injection. Expression of the beta-galactosidase gene product was monitored by polymerase chain reaction and histochemistry. RESULTS: Up to 28 days after in vivo gene transfer, beta-galactosidase activity could be demonstrated in the pancreas. Intraductal application induced gene expression in lining duct cells preferentially. Twenty-four hours after intraductal injection of liposomes, a dose-dependent, transient increase in serum amylase levels was detected. Nevertheless, no histological signs of pancreatitis were evident. Intra-arterial injection resulted in beta-galactosidase expression in endothelial cells of intrapancreatic arteries, as well as in the spleen, lymph nodes and liver, but not in ductal cells of the pancreas. Only occasionally were acinar cells positive for blue staining by either type of treatment. CONCLUSION: These experiments demonstrate that in vivo gene transfer into the pancreas is feasible using DNA-liposome complexes. Furthermore, the route of administration largely determines cell type specificity and side-effects. This technique might have an impact for the development of gene therapy strategies for pancreatic diseases.

Liposome mediated in vivo gene transfer into different tissues of the gastrointestinal tract.

The possibility to transfer and express genetic material in mammalian cells represents a new approach to the treatment of genetic and acquired disorders. So far, most studies use in vitro techniques to introduce foreign DNA into cultured cells, followed by reintroduction of these genetically altered cells into living organisms. In the present study we demonstrate that the LacZ marker gene can be selectively delivered, by in vivo techniques, to various locations of the gastrointestinal tract. Genetic material was targeted to the stomach, the colon, the liver and the pancreas using cationic liposomes. For transfer into the stomach and colon an intraluminal application, in the liver a portal access and in the pancreas an intraductal infusion was chosen. 48 hours after administration, the LacZ gene product beta-galactosidase could be localized in these tissues by cytochemistry. These experiments suggest a new approach to study gastrointestinal physiology and may offer novel aspects for the treatment of gastrointestinal diseases.