Intron 1 retaining cyclooxygenase 1 splice variant is induced by osmotic stress in human intestinal epithelial cells.

The biological roles of intron 1 retaining cyclooxygenase (Cox) 1 splice variants Cox-3 and PCox-1a (Cox-1ir) are not known. In humans, Cox-3 transcription has previously been shown to occur in the brain and in the aorta. However, conclusive evidence regarding the existence of a human Cox-3 protein is lacking. We studied the expression of intron 1 retaining cyclooxygenase 1 splice variants in the human colon cancer cell line Caco-2 and in human colonic tissue samples. In Caco-2 cells, their transcription was induced up to 47-fold by osmotic stress. The corresponding protein, however, could not be detected by Western blotting. In human colonic tissue samples derived from intact and inflamed areas, a low level of Cox-1ir mRNA (1500 +/- 1280 copies per 100 ng total RNA; mean+/-standard deviation; n = 20) was also found. In Caco-2 cells, induction of Cox-1ir under osmotic stress was reversed by addition of the organic osmolyte betaine. Under hypertonic but not under isotonic conditions, splice variant-specific degradation of Cox-1ir mRNA using RNA interference resulted in increased production of fully spliced Cox-1 and Cox-2 mRNA (P = 0.002). In summary, our results indicate that the intron 1 retaining Cox-1 splice variant RNA molecules are expressed by human intestinal epithelial cells in a controlled manner, are most likely not translated and play a regulatory role in the cyclooxygenase mediated epithelial osmoregulation.

Severe acute pancreatitis: prognostic factors in 270 consecutive patients.

Acute pancreatitis (AP) is a common abdominal disorder with severity varying from mild to fatal disease. Predicting a patient's outcome remains problematic. The aim of this study was to analyze a large consecutive series of patients with severe AP and to identify prognostic factors for hospital mortality. Between 1989 and 1997, a consecutive series of 270 patients with severe AP were included in the study. All patients fulfilled the criteria of Atlanta classification for severe AP. Retrospectively and prospectively collected data included age, gender, etiology, number of previous episodes of pancreatitis, medication history, type of admission, body-mass index (BMI), respiratory failure, renal failure, need for pressor support, and abdominal surgery performed during hospitalization. The overall mortality rate was 24.4%. In univariate survival analysis advanced age, history of continuous medication, patient transferred from other hospital, high BMI, respiratory or renal failure, need for pressor support, and need for abdominal surgery were significant prognostic factors for hospital mortality. In a multivariate stepwise logistic regression analysis, the need of pressor support, renal failure requiring dialysis, advanced age, history of continuous medication and need for abdominal surgery were identified as independent prognostic factors for mortality. A logistic regression analysis of variables available on admission (the first seven above mentioned variables) showed that transferral admission, advanced age, and history of continuous medication were independent prognostic factors for mortality. In patients with severe AP, advanced age, history of continuous medication, and need for dialysis, mechanical ventilator support, and pressor support predict fatal outcome and thus should be taken into account in clinical evaluation.

Advances in the laboratory diagnostics of acute pancreatitis.

Acute pancreatitis is a rather common abdominal disorder. In most patients the disease is mild, but about 20% of cases develop a severe necrotizing form of the disease with complications. In an emergency setting, the diagnosis of acute pancreatitis remains problematic and several patients with severe disease are diagnosed only at autopsy. Measurements of amylase or lipase are the principal laboratory methods for diagnosing acute pancreatitis. However, their sensitivity and specificity are generally considered unsatisfactory. Recent advances in the knowledge of the pathogenesis of acute pancreatitis and advances in laboratory technology have revealed new diagnostic possibilities. Especially assays based on trypsin pathophysiology have brought new alternatives for diagnostics and severity grading of the disease. Additionally, development of phospholipase A2 determinations and discovery of a new pancreatic protein, pancreatitis-associated protein, are very interesting. This article summarizes the value of new methods in the laboratory diagnostics of acute pancreatitis.

Rapid measurement of urinary trypsinogen-2 as a screening test for acute pancreatitis.

BACKGROUND: Acute pancreatitis can be difficult to diagnose. We developed a rapid dipstick screening test for pancreatitis, based on the immunochromatographic measurement of urinary trypsinogen-2. METHODS: We prospectively compared the urinary trypsinogen-2 dipstick test with a quantitative urinary trypsinogen-2 assay, a urinary dipstick test for amylase, and serum and urinary amylase assays in 500 consecutive patients with acute abdominal pain at two emergency departments. Acute pancreatitis was diagnosed according to standardized criteria. RESULTS: The urinary trypsinogen-2 dipstick test was positive in 50 of the 53 patients with acute pancreatitis (sensitivity, 94 percent), including all 7 with severe pancreatitis. Two patients with urinary trypsinogen-2 concentrations below the sensitivity threshold of the test (50 ng per milliliter) and one with a very high concentration had false negative results. The test was also positive in 21 of the 447 patients without pancreatitis (specificity, 95 percent), including 7 with abdominal cancers, 3 with cholangitis, and 2 with chronic pancreatitis. The sensitivity and specificity of the dipstick test were similar to those of the quantitative urinary trypsinogen-2 assay and higher than those of the urinary amylase dipstick test. The serum amylase assay had a sensitivity of 85 percent (with a cutoff value of 300 U per liter for the upper reference limit) and a specificity of 91 percent. The sensitivity and specificity of the urinary amylase assay (cutoff value, 2000 U per liter) were 83 and 88 percent, respectively. CONCLUSIONS: In patients with acute abdominal pain seen in the emergency department, a negative dipstick test for urinary trypsinogen-2 rules out acute pancreatitis with a high degree of probability. A positive test usually identifies patients in need of further evaluation.

Acceleration of wound healing in aged rats by topical application of transforming growth factor-beta(1).

The impaired wound healing associated with aging may reflect inadequate secretion or delivery of cytokines. Transforming growth factor-beta(1) is a mitogenic polypeptide with beneficial effects on wound healing. In the present study we questioned whether topical administration of transforming growth factor-beta(1) could improve the wound healing process in aged rats in vivo. Wound repair (from 1 to 14 days) was analyzed in full-thickness incisional wounds from 2-year-old rats with or without a single topical application of transforming growth factor-beta(1) (1 microg/wound) at the time of wounding. Identical wounds from 3-month-old, untreated rats served as controls. Histologic analysis showed a marked delay in several aspects of wound repair in the aged rats in comparison with that noted in the younger animals. Immunostaining of the wounds for proliferating cell nuclear antigen showed a reduction in the number of cycling fibroblasts in old rats. In addition, the number of capillaries per unit area of the wound as determined by a stain for Griffonin (Bandeiraea) simplicifolia lectin, and the number of inflammatory cells as identified by an antibody specific for macrophages, were also reduced in the wound area in old rats. Treatment with transforming growth factor-beta(1) resulted in marked enhancement of the following parameters: cell proliferation, inflammatory cell and fibroblast influx, wound closure, and angiogenesis. As seen with in situ hybridization, a similar temporal pattern of expression of messenger RNAs corresponding to type I procollagen and Secreted Protein, Acidic and Rich in Cysteine (osteonectin), known to be prevalent in healing wounds, was observed in both young and aged rats. However, the levels of mRNA corresponding to these secreted proteins appeared to be reduced in wound tissue from aged rats. Treatment with transforming growth factor-beta(1) subsequently resulted in an increase in the expression of both type I procollagen and Secreted Protein, Acidic and Rich in Cysteine mRNA in the wound tissue from aged rats. In summary, a single topical application of transforming growth factor-beta(1) to the wounds of aged rats at the time of wounding was associated with a healing response that, in all the parameters of wound repair examined, was similar to that of young rats. Topical transforming growth factor-beta(1) might therefore be beneficial in the treatment of dermal wounds in the aged.

Novel delivery system for inducing quiescence in intestinal stem cells in rats by transforming growth factor beta 1.

BACKGROUND/AIMS: Intestinal mucosa, a tissue in a dynamic state of rapid cellular proliferation, is often adversely affected by cytotoxic drugs. The purpose of this study was to develop an oral delivery system targeting transforming growth factor (TGF) beta 1 locally and analyze its effects on the epithelial stem cells of gastrointestinal mucosa. METHODS: Rats were treated with recombinant TGF-beta 1 in alginate beads perorally or with recombinant TGF-beta 1 in phosphate-buffered saline perorally or intraperitoneally. Control animals received phosphate-buffered saline only. The size of the villi was measured. Proliferating and mitotic indices were determined by quantifying immunohistochemical staining for proliferating cell nuclear antigen. RESULTS: Alginate beads released no TGF-beta 1 in acid. However, in pH 7.4, TGF-beta 1 was released in an active form. Histomorphometrical analysis showed a marked reduction in villus height (50%-70%) in the intestinal mucosa of animals treated perorally with recombinant TGF-beta 1 in alginate beads. Also, the proliferating and mitotic indices were significantly reduced (P < 0.01) in these animals as compared with controls and other routes of administration. CONCLUSIONS: This study shows that recombinant TGF-beta 1 administered using a novel oral delivery system induces stem cell quiescence in the intestinal mucosa of the rat.

Fatal air embolism as a complication of laser-induced hyperthermia.

Laser-induced hyperthermia is a new treatment modality for malignancies. When large tumors are treated with laser thermia, relatively high laser powers (4-6 W) must be used to accomplish destruction of the cancerous cells. Overheating and burning of the laser fibers and contact tips during laser thermia treatment must be prevented by using coaxial gas or fluid flow. The present report describes a case of fatal air embolism as a complication of laser thermia using coaxial gas. The authors warn all surgeons performing this procedure not to use coaxial air flow for cooling the contact tips because of the potential risk of air embolism.