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Efficacy of silver-coating central venous catheters in reducing bacterial colonization.

OBJECTIVE: To compare silver-coated and uncoated central venous catheters regarding bacterial colonization. To assess the relative contribution of catheter hub and skin colonization to catheter tip colonization. DESIGN: Prospective, randomized clinical trial. SETTING: Intensive care unit in a university hospital. PATIENTS: Patients after cardiac surgery who required a central venous double-lumen catheter (DLC). INTERVENTIONS: Sixty-seven adult patients were prospectively randomized to receive either a silver-coated (S group, n = 34) or an uncoated control (C group, n = 33) DLC. Blood cultures were drawn at catheter removal, and removed catheters were analyzed with quantitative cultures. Typing of microorganisms included DNA fingerprinting. MEASUREMENTS AND MAIN RESULTS: Catheters were removed if no longer necessary and aseptically divided into three segments: segment A, the catheter tip; segment B, an intermediate section; and segment C, the subcutaneous portion. Bacterial catheter colonization was quantitatively measured using sonication to detach adherent bacteria from the catheter segments in the broth and subsequent culture of an aliquot. Selected isolates of coagulase-negative staphylococci and other bacteria from catheter segments were examined by means of pulsed-field gel electrophoresis (PFGE) after macrorestriction digestion of bacterial DNA to study colonization pathogenesis. Quantitatively lower bacterial colonization could be demonstrated on the silver-coated catheters (200 +/- 550 colony forming units [CFUs]/cm catheter segment; mean +/- SD). The difference in the control catheters (1120 +/- 5350 CFUs/cm catheter segment; mean +/- SD) was not, however, significant (p = .25). The frequency of colonization of at least one catheter segment was 52.9% for the silver-coated catheters and 57.6% for the control catheters (p= .44), without any significant differences in the colonization of corresponding catheter segments. The rate of significant catheter colonization (i.e., > or = 10(3) CFUs/cm catheter by quantitative catheter culture or > or = 10(3) CFUs/mL by luminal flush) was nine in the silver group and seven in the control group, a difference that failed to reach significance (p = .41). Two patients in both groups developed catheter-related bacteremia. Pattern analysis after PFGE demonstrated that about 70% of the isolates found on the catheter tip were identical with those on the skin at the insertion site, whereas about 75% were identical with those recovered from the hub. In 29% of colonized catheters, identical bacteria were found on the hub and the skin at the insertion site. CONCLUSIONS: Silver-coating of DLCs did not significantly reduce bacterial catheter colonization compared with the control catheters. PFGE analysis of coagulase-negative staphylococci and other bacteria demonstrated various pathogenic routes of catheter-related colonization, whereby the microorganisms of the skin flora around the insertion site must be regarded as the main source of catheter-related infections.

Adult↗

Neonatal airway colonization with gram-negative bacilli: association with severity of bronchopulmonary dysplasia.

BACKGROUND: Airway colonization with Gram-negative bacilli (GNB) and Gram-positive cocci (GPC) is common in mechanically ventilated neonates. Whether GNB are related to nosocomial bloodstream infection (BSI) and/or to the severity of bronchopulmonary dysplasia (BPD) is unknown. METHODS: We prospectively examine this relationship using a cohort design. Data from 260 < or = 1250-g birth weight inborn infants (1991 to 1995) intubated > or = 2 weeks included 917 serial tracheal cultures and 583 blood cultures. The severity of BPD was assessed by duration of mechanical ventilation, oxygen dependency at 36 weeks of postconceptional age and the use of home oxygen supplementation. RESULTS: After 2 weeks of ventilation, 80% of the infants were colonized with GPC (Staphylococus epidermidis and Staphylococcus haemolyticus in 90% of the cases). Superimposed on 36% of these infants was GNB airway colonization with Klebsiella pneumoniae (25%), Enterobacter cloacae (25%), Escherichia coli (25%), Pseudomonas aeruginosa (10%), Serratia marcescen (10%), Acinetobacter baumannii and Haemophilus influenzae (5%). Comparison between 174 GPC- and 86 GNB-colonized infants showed that demographics, birth weight, gestational age, perinatal risk factors and mortality were similar. Fifteen percent of GNB-colonized infants developed BSI caused by GNB and 14% developed BSI caused by GPC. No significant temporal relationship between airway colonization and BSI was noted. GNB infants were ventilated longer and required oxygen at 36 weeks of postconceptional age and home oxygen supplementation twice as often as infants colonized only with GPC. GNB colonization was a predictor of severe BPD after controlling for ventilation. Ureaplasma colonization occurred in 28% of GNB-colonized and 33% of noncolonized infants and was not a predictor of BPD severity. CONCLUSION: GNB airway colonization creates a moderate risk for BSI. Antibiotic treatment does not regularly eradicate GNB. GNB airway colonization is associated with severe BPD, but further studies will be necessary before therapeutic efforts to eradicate GNB from the airways should be undertaken.

Bronchopulmonary Dysplasia↗

Predictors for Haemophilus influenzae colonization, antibiotic resistance and for sharing an identical isolate among children attending 16 licensed day-care centers in Michigan.

BACKGROUND: Nontypable Haemophilus influenzae is an important cause of otitis media in children. Children attending day-care centers are at an increased risk for nontypable H. influenzae colonization and otitis media. We describe the prevalence of nontypable H. influenzae colonization, antibiotic resistance and predictors for colonization and sharing an identical isolate with at least 1 other child in the same day-care centers among children attending 16 day-care centers. METHODS: Throat swabs of 198 children < 3 years old attending 16 day-care centers were cultured for H. influenzae. Day-care center directors and parents completed risk factors questionnaires. Nontypable H. influenzae isolates were screened for antibiotic resistance and genotyped. Statistics were performed using SAS software (SAS Institute, Inc., Cary, NC). RESULTS: We isolated 179 unique nontypable H. influenzae strains from 127 participants. Colonization ranged from 0% to 95% among day-care centers. As individual factors, exposure to tobacco smoke was associated with colonization (P = 0.05), and racial self-identifications as "other" (nonwhite, nonblack) was protective (P = 0.035), whereas as "black" was protective for sharing (P = 0.03). Pacifier use was associated with sharing (P = 0.04), but not with colonization. As day-care centers factors, rates of colonization and sharing were higher in day-care centers with > or = 5 classrooms (P < 0.01 and P = 0.03), with such suboptimal hygiene habits as minimal hand washing by staff after eating (P < 0.002 and P < 0.01) or by children after wiping their own nose (P = 0.01 and P = 0.003). Of colonized children, 41% presented a beta-lactamase-producing strain. Colonized children were more likely to carry resistant strains if they were taking an antibiotic (P = 0.02). CONCLUSION: Although day-care center colonization varied, the overall colonization rate was high. Colonization with nontypable H. influenzae, with beta-lactamase-producing strain and sharing were, mostly, associated with modifiable risk factors.

Carrier State↗

The impact of colon preparation timing on colonoscopic detection of colorectal neoplasms--a prospective endoscopist-blinded randomized trial.

OBJECTIVES: Timing of colon preparation might influence the cleansing condition of the colon and therefore affect the quality and diagnostic yield of colonoscopy. This study compared two different timings of colon preparation to assess the efficacy of colon cleansing and diagnostic yield of colorectal neoplasms during colonoscopy. METHODS: One hundred twenty-one eligible subjects who had colorectal neoplasms detected at a screening colonoscopy were randomly assigned to receive colon preparation with polyethylene glycol electrolyte lavage solution (PEG-ELS) either on the day of (on the day group) or the night before (previous night group) a second colonoscopy. The condition of colon preparation and the diagnostic yield of colorectal neoplasms were recorded and compared between the two groups for the initial and second colonoscopies. RESULTS: Fifty-nine subjects received colon preparation on the night before and 60 subjects received colon preparation the day of a second colonoscopy. Colon preparation for the second colonoscopy was better in the on-the-day group than in the previous night group (P= 0.003). Colon preparation for the second colonoscopy was better for each group in comparison with the initial colonoscopy (P < 0.0001). An interobserver variability analysis using 20 randomly selected colonoscopies from the two groups revealed good correlation among four colonoscopists. More lesions were detected in group A during the second colonoscopy (P= 0.028). CONCLUSION: Colon preparation on the day of colonoscopy has a better cleansing quality and higher diagnostic yield. Subjects who had lesions detected during the initial screening colonoscopy had better colon cleansing for the second colonoscopy regardless of the timing of preparation.

Aged↗

Adaptation of Campylobacter jejuni NCTC11168 to high-level colonization of the avian gastrointestinal tract.

The genome sequence of the human pathogen Campylobacter jejuni NCTC11168 has been determined recently, but studies on colonization and persistence in chickens have been limited due to reports that this strain is a poor colonizer. Experimental colonization and persistence studies were carried out with C. jejuni NCTC11168 by using 2-week-old Light Sussex chickens possessing an acquired natural gut flora. After inoculation, NCTC11168 initially colonized the intestine poorly. However, after 5 weeks we observed adaptation to high-level colonization, which was maintained after in vitro passage. The adapted strain exhibited greatly increased motility. A second strain, C. jejuni 11168H, which had been selected under in vitro conditions for increased motility (A. V. Karlyshev, D. Linton, N. A. Gregson, and B. W. Wren, Microbiology 148:473-480, 2002), also showed high-level intestinal colonization. The levels of colonization were equivalent to those of six other strains, assessed under the same conditions. There were four mutations in C. jejuni 11168H that reduced colonization; maf5, flaA (motility and flagellation), and kpsM (capsule deficiency) eliminated colonization, whereas pglH (general glycosylation system deficient) reduced but did not eliminate colonization. This study showed that there was colonization of the avian intestinal tract by a Campylobacter strain having a known genome sequence, and it provides a model for colonization and persistence studies with specific mutations.

Adaptation, Biological↗

Immunization with Staphylococcus aureus clumping factor B, a major determinant in nasal carriage, reduces nasal colonization in a murine model.

Staphylococcus aureus is responsible for a wide range of infections, including soft tissue infections and potentially fatal bacteremias. The primary niche for S. aureus in humans is the nares, and nasal carriage is a documented risk factor for staphylococcal infection. Previous studies with rodent models of nasal colonization have implicated capsule and teichoic acid as staphylococcal surface factors that promote colonization. In this study, a mouse model of nasal colonization was utilized to demonstrate that S. aureus mutants that lack clumping factor A, collagen binding protein, fibronectin binding proteins A and B, polysaccharide intercellular adhesin, or the accessory gene regulator colonized as well as wild-type strains colonized. In contrast, mutants deficient in sortase A or clumping factor B (ClfB) showed reduced nasal colonization. Mice immunized intranasally with killed S. aureus cells showed reduced nasal colonization compared with control animals. Likewise, mice that were immunized systemically or intranasally with a recombinant vaccine composed of domain A of ClfB exhibited lower levels of colonization than control animals exhibited. A ClfB monoclonal antibody (MAb) inhibited S. aureus binding to mouse cytokeratin 10. Passive immunization of mice with this MAb resulted in reduced nasal colonization compared with the colonization observed after immunization with an isotype-matched control antibody. The mouse immunization studies demonstrate that ClfB is an attractive component for inclusion in a vaccine to reduce S. aureus nasal colonization in humans, which in turn may diminish the risk of staphylococcal infection. As targets for vaccine development and antimicrobial intervention are assessed, rodent nasal colonization models may be invaluable.

Adhesins, Bacterial↗

Severe imbalance of cell proliferation and apoptosis in the left colon and in the rectosigmoid tract in subjects with a history of large adenomas.

BACKGROUND: Alterations in epithelial proliferation and apoptosis in colonic mucosa are associated with an increased risk of colon cancer. It is unclear if these alterations represent a generalised "field defect". AIMS: To analyse segmental patterns of cell proliferation and apoptosis in the colon of subjects with a high and no apparent risk of colon cancer. METHODS: Pancolonoscopy was performed in 15 patients with resected adenomas (> or =1.5 cm) and in nine subjects without an apparent risk of colorectal cancer. Mucosal biopsies were taken from the right colon, left colon, and sigmoid rectum. Crypt cell proliferation and apoptosis were evaluated, respectively, with bromodeoxyuridine immunohistochemistry and terminal deoxyuridine nucleotidyl nick end labelling of DNA strand breaks. Results are expressed as total labelling index (TLI) and labelling index (LI) for each of the five compartments in which colonic crypts were divided (fourth and fifth compartments were evaluated together) for cell proliferation and as apoptotic index (AI) for apoptosis assessment. RESULTS: No significant segmental variations in proliferation were found in either group. Compared with controls, adenoma patients had higher TLIs for the right (p>0.05), left (p<0.005), and sigmoid rectum (p<0.05) segments, and higher left colon LIs for crypt compartments (compartment 1, p<0.01; compartment 2, p<0.005; compartment 3, p<0.001; compartments 4-5, p<0.01). Control AIs were similar in all segments but in the adenoma patients left colon and sigmoid rectum AIs were lower than their right colon indexes (p<0.05, p<0.05) and corresponding values for controls (p<0.01, p<0.05). CONCLUSIONS: The colonic mucosa of patients with past adenomas presents diffuse hyperproliferation and, distally, abnormally distributed proliferating cells and markedly reduced apoptosis. These changes represent a significant risk for malignancies and could account for the high prevalence of left colon tumours.

Adenoma↗

Risk factors for pneumonia, and colonization of respiratory tract and stomach in mechanically ventilated ICU patients.

Risk factors for the development of ventilator-associated pneumonia (VAP) and colonization of the respiratory tract and stomach with enteric gram-negative bacteria (EGB) and Pseudomonadaceae were determined in 141 ventilated patients using univariate analysis and the Cox proportional hazards model. VAP was caused by EGB in 14 patients (10%), and by Pseudomonadaceae in 19 patients (13%). The duration of ventilation was a significant risk factor for VAP caused by EGB and Pseudomonadaceae, and for acquired colonization in oropharynx, stomach, and trachea with these species. Of 20 other variables, oropharyngeal colonization with EGB on admission (hazard ratio [HR] = 4.5) and an infection on admission (HR = 2.7) were selected as risk factors for VAP caused by EGB. Acquired colonization with Pseudomonadaceae in oropharynx (HR = 5.0) was the most important risk factor for VAP caused by these species. Gastric colonization with EGB or Pseudomonadaceae were no risk factors for VAP. For acquired oropharyngeal colonization with EGB only the duration of ventilation was a risk factor, whereas preceding colonization of the trachea with Pseudomonodaceae and duration of ventilation were risk factors for acquired oropharyngeal colonization with these species. In the Cox model, only the duration of ventilation was significantly related to acquired gastric colonization with EGB. Preceding colonization of the orophayrnx and of the trachea with Pseudomonadaceae were risk factors for acquired colonization with these species in the stomach. Twelve patients with VAP (46%) and 38 without VAP (33%) died (p = 0.21). In conclusion, duration of ventilation and colonization of the upper respiratory tract are the most important risk factors for VAP caused by EGB or Pseudomonadaceae.

Adult↗

Role of different routes of tracheal colonization in the development of pneumonia in patients receiving mechanical ventilation.

STUDY OBJECTIVE: To evaluate the importance of the different pathogenic pathways involved in the development of ventilator-associated pneumonia (VAP). DESIGN: Prospective study. SETTING: An 18-bed medical and surgical ICU. PATIENTS: One hundred twenty-three patients receiving mechanical ventilation (MV). INTERVENTIONS: Tracheal, pharyngeal, and gastric samples were obtained simultaneously every 24 h. In cases where VAP was suspected clinically, bronchoscopy with protected specimen brush and BAL were performed. Semiquantitative cultures of pharyngeal samples and quantitative cultures for the remaining samples were obtained. RESULTS: Tracheal colonization at some time during MV was observed in 110 patients (89%). Eighty patients had initial colonization, 34 patients had primary colonization, and 50 patients had secondary colonization. Nineteen patients had VAP, and 25 organisms were isolated. For none of these organisms was the stomach the initial site of colonization. Gram-positive organisms colonized mainly in the trachea during the first 24 h of MV (p<0.001). On the contrary, enteric Gram-negative bacilli (p<0.001) and yeasts (p<0.002) colonized the trachea secondarily. Previous endotracheal intubation (p<0.005) and acute renal failure before admission to the ICU (p<0.001) were associated with colonization by Pseudomonas aeruginosa; prior antibiotics were associated with colonization by Acinetobacter baumanii (p<0.05) and yeasts (p<0.006); and cranial trauma was associated with Staphylococcus aureus colonization (p<0.035). CONCLUSIONS: Although the stomach can be a source of organisms that colonize the tracheobronchial tree, it is a much less common source of the bacteria that cause VAP. The pattern of colonization and risk factors may be different according to the type of organisms involved.

Adolescent↗

Prophylactic fluconazole is effective in preventing fungal colonization and fungal systemic infections in preterm neonates: a single-center, 6-year, retrospective cohort study.

OBJECTIVE: Despite the promising preliminary results observed in extremely low birth weight (ELBW) populations, the use of fluconazole to prevent fungal colonization and infection in preterm neonates in the NICU is still an open question and not yet recommended as a standard of care. We have reviewed our 6-year series to assess the effectiveness and safety of this form of prophylaxis. METHODS: This retrospective study consisted of 465 neonates who weighed < 1500 g at birth and were admitted to our NICU in the period 1998-2003. Those who were born between 1998 and 2000 and did not receive fluconazole prophylaxis (group A, n = 240) were compared with those who were born between 2001 and 2003 and treated with fluconazole until the 30th day of life (45th for neonates < 1000 g at birth; group B, n = 225). Weekly surveillance cultures were obtained from all patients. Incidence of fungal colonization, incidence of systemic fungal infection (SFI), rate of progression from colonization to infection, and mortality rates attributable to fungi were calculated for both groups and separately for neonates who were < 1000 g (ELBW) and were 1001 to 1500 g (NE-VLBW) at birth. RESULTS: Overall fungal colonization was significantly lower in group B (24.0%) than in group A (43.8%; relative risk [RR]: 0.406; 95% confidence interval [CI]: 0.273-0.605). The same was true of neonates with colonization in multiple sites (2.6% vs 5.8%) and of those with colonization from high-risk sites (5.8% vs 19.2%). SFI incidence was significantly lower in group B (10 of 225 cases; 4.4%) than in group A (40 of 240 cases; 16.7%; RR: 0.233; 95% CI: 0.113-0.447). Reduction of both colonization and SFI in group B was greater in the ELBW neonates and also significant in the NE-VLBW neonates. Rate of progression from colonization to infection was significantly lower in group B (0.17 vs 0.38; RR: 0.369; 95% CI: 0.159-0.815). Crude mortality rate attributable to Candida species was 1.7% (4 of 240) in group A vs 0% (0 of 225) in group B. Overall mortality rate (any cause before hospital discharge) was similar in the two groups (11.2% vs 10.6%), but in colonized infants (n = 159), it was significantly lower in group B (3.7% vs 18.1%; RR: 0.174; 95% CI: 0.039-0.778). The incidence of natively fluconazole-resistant fungal species did not increase over the years, and patterns of sensitivity to fluconazole remained the same. No adverse reaction related to fluconazole occurred. CONCLUSIONS: Prophylactic fluconazole significantly reduces the incidence of colonization and systemic infection by Candida species in both ELBW and NE-VLBW neonates and decreases the rates of progression from initial colonization to massive colonization and to systemic infection. All VLBW neonates may benefit from fluconazole prophylaxis.

Antifungal Agents↗

Methicillin-resistant Staphylococcus aureus colonization and its association with infection among infants hospitalized in neonatal intensive care units.

OBJECTIVES: We conducted this study to assess the rate of methicillin-resistant Staphylococcus aureus colonization and its association with infection among infants hospitalized in methicillin-resistant S aureus-endemic NICUs. METHODS: Between March 2003 and February 2004, surveillance culture specimens from the nares, postauricular areas, axillae, and umbilicus of infants admitted to the NICUs at a children's hospital in Taiwan were obtained weekly for the detection of methicillin-resistant S aureus. All colonized and clinical isolates from each study infant with methicillin-resistant S aureus infection were genotyped with pulsed-field gel electrophoresis, with Sma1 digestion, and compared. RESULTS: A total of 783 infants were included in this study. Methicillin-resistant S aureus colonization was detected for 323 infants during their NICU stays, with detection with the first 2 samples for 89%. Nares and umbilicus were the 2 most common sites of initial colonization. Methicillin-resistant S aureus colonization was associated significantly with premature birth (< or = 28 weeks) and low birth weight (< or = 1500 g), and infants with colonization had a significantly higher rate of methicillin-resistant S aureus infection, compared with those without colonization (26% vs 2%). Methicillin-resistant S aureus colonization was noted for 84 of 92 infants with methicillin-resistant S aureus infections. Of the 68 episodes with previous colonization and isolates available for genotyping analysis, colonized and clinical isolates were indistinguishable in 63 episodes, highly related in 2 episodes, and distinct in 3 episodes. CONCLUSIONS: More than 40% of the hospitalized infants were colonized with methicillin-resistant S aureus during their stay in methicillin-resistant S aureus-endemic NICUs; this was associated significantly with methicillin-resistant S aureus infection. Most infants with methicillin-resistant S aureus infections had previous colonization with an indistinguishable strain.

Axilla↗

Colonization of suckling pigs by Streptococcus suis with particular reference to pathogenic serotype 2 strains.

Three swine commercial farms with high mortality rates in nursery pigs due to Streptococcus suis serotype 2 were studied. Brain samples from diseased animals were collected for a period of 6 to 10 mo and used to isolate the strain that was responsible for the mortality (virulent strain) in each farm. Tonsil swabs from piglets at 5, 10 and 15 d were taken to assess both total colonization and colonization by the virulent strain. The effect of sow vaccination against S. suis on colonization was evaluated in 1 of the farms. All suspect tonsil isolates were identified biochemically and then tested against serotype 2. The genomic patterns of serotype 2 isolates were compared to that of the virulent strain using Rep-PCR. Results showed that total colonization by S. suis occurred very early in the pigs' life, with most animals being colonized by weaning age. Prevalence of colonization by serotype 2 strains was much lower than total colonization. After comparing serotype 2 isolates with the virulent strains, only 1 tonsillar isolate had the same genomic pattern as the virulent strain and it belonged to a 4-week-old weaned pig. The genomic pattern of the virulent strain was not found in any tonsillar isolate from 15-day-old or younger pigs. Although limited by sample size, sow vaccination against S. suis increased total colonization at the same time significantly decreasing colonization by serotype 2 strains. Even though most pigs are colonized early in age by S. suis, colonization by the virulent strain is of low prevalence and delayed in time. This could constitute a risk factor for developing the disease later in time, because animals would be colonized when maternal immunity is no longer present, allowing the organism to become systemic.

Animals↗

Long-term follow-up after resectional surgery in patients with Crohn's disease involving the colon.

BACKGROUND: The majority of patients with Crohn's disease will eventually have colonic involvement, and more than 50% of these patients undergo resectional colonic surgery. The extent of colonic resection is discussed controversially. AIMS: We evaluated prognostic factors influencing the long-term outcome after resectional surgery including the colon. METHODS: We analyzed the postoperative course in 170 patients (mean follow-up 7.4 years) after first colonic surgery. Lifetable and multivariate factor analysis were performed to assess the influence of various factors on the postoperative long-term outcome. 85% of the patients had concomitant ileal disease, 40% had rectal disease, 48% percent of the patients had extensive colonic disease at the time of primary surgery. RESULTS: In 17% of the initial operations a colectomy was performed, the remaining 83% operations consisted in segmental colonic resections. The cumulative risks of clinical recurrence/reoperation were 63%/33% after ten years and increased by the presence of anal fistulas (relative risk 1.7/3.0) and after colocolonic type of anastomosis (relative risk 1.9/2.8). Ileal disease, rectal disease, extent of resection and pattern of colitis did not influence the recurrence rates. The risk to undergo completion colectomy was 11% ten years after segmental resection and not higher in the presence of extensive colonic disease. The risk of a definitive stoma was 11% after ten years and higher after ileorectal anastomosis (25% versus 8% after segmental resection: p < 0.003). CONCLUSION: Colocolonic type of anastomosis and the presence of anal fistulas are risk factors for recurrence after initial colonic resection. Segmental resections were not followed by increased recurrence rates or a higher stoma rate. To maintain colonic length and intestinal continuity segmental colonic resection is the treatment of choice in patients undergoing surgery for local complications, even in the presence of extensive colonic disease.

Adult↗

The effects of long-term ingestion of asbestos on the colon of F344 rats.

Weanling F344 rats, which were fed a diet containing 10% chrysotile (B), were studied over their life-time to determine the effects of ingested asbestos on the colon. Control groups consisted of rats fed a diet containing a 10% nonnutritive cellulose or a standard laboratory rat diet. The pathological findings in the colons of 501 rats (189 on asbestos diet, 197 on fiber control diet, and 115 on standard control diet), are reported here. Epithelial tumors of the colon (eight adenocarcinomas and one adenoma) were found in nine of the rats on study. Four of the tumors were in asbestos-fed rats, two tumors were found in the non-nutritive cellulose controls, and three tumors were found in the standard laboratory rat diet controls. The probability (based on actuarial analysis) of developing adenoma or adenocarcinomas during the 32 months of the study were 7.4% for the asbestos-fed group, 3.5% for the fiber control diet and 4.0% on the standard control diet. In addition, one malignant mesothelioma of the type induced by intraperitoneally administered asbestos was found in the asbestos-fed group. Non-neoplastic lesions of the colon were also evaluated. The cumulative risk for development of any colon-associated lesion (non-neoplastic plus neoplastic lesions) was greatest for asbestos-fed rats (17.9%), compared to 13.6% for those fed the fiber control diet and 8.2% for those fed the standard control diet. The colon tissue levels of adenosine, 3'-5'-cyclic monophosphate (cAMP) were significantly lower in the animals fed asbestos compared to the control diets. Chrysotile fibers were seen by electronmicroscopy (e.m.) in six of ten ashed colon specimens of rats fed the asbestos diet. Although the differences in numbers of tumors between the animals fed asbestos and the controls were not statistically significant at the 5% level, we felt that the combination of observations including 1) evidence of increased probability of asbestos-fed animals to develop colon lesions in general; 2) evidence of a special type of mesothelioma in rats fed asbestos; 3) evidence for a cell regulator defect (lowered cAMP levels) in colon tissues of animals fed asbestos; and 4) evidence for asbestos fiber penetration of the colonic mucosa (e.m. studies) suggest that ingested asbestos is not inert in the colon.

Adenocarcinoma↗

Insulin binding by normal and neoplastic colon tissue.

Insulin receptors in human colon tumours, normal colon tissue and mesenteric fat removed at surgery have been identified by measuring the binding of labelled insulin to cell membrane preparations. Insulin binding sites were readily detected in all tissues, with mean +/- SD binding site concentrations of 43 +/- 41, 44 +/- 39, and 44 +/- 35fmol/mg membrane protein, and dissociation constants of 0.73 +/- 0.61, 0.66 +/- 0.41, and 0.78 +/- 0.58nM for microsomal plasma membrane preparations of tumour (n = 23) respectively. The specificity of binding of labelled insulin was similar in tumour and normal colon samples. Binding in normal colon preparations was highest in the epithelium (84fmol/mg membrane protein) and lower in lamina propria (19fmol/mg), submucosa (25fmol/mg) and muscle wall (13fmol/mg). Degradation of labelled insulin was similar in tumour and normal colon preparations. Mean receptor levels were not significantly different between microsomal membrane preparations and plasma membranes partially purified on discontinuous sucrose gradients, quantitated against either unit membrane protein or unit 5'-nucleotidase specific activity. There was a significant negative correlation between insulin levels, but no significant relationship was seen between serum insulin and receptor levels in either colon tumour or tissue preparations from full-thickness normal colon wall. An inverse correlation between serum insulin and receptor levels was, however, apparent in preparations of colonic musosa. These data indicate that although insulin receptors in colon tumours share the same biochemical characteristics as those present in the normal colon, receptors in tumour tissue are less sensitive to down-regulation by ambient insulin than receptors in mesenteric fat cells and normal colonic mucosa.

Adipose Tissue↗

Expression of urokinase-type plasminogen activator and plasminogen activator inhibitor in colon disease.

Surveillance colonoscopy and biopsy are inaccurate methods of predicting the likelihood of ulcerative colitis patients to develop colon carcinoma. We examined uPA and PAI-1 as potential markers for assessing these patients and those with familial polyposis who are at risk of developing colon cancer. For comparison, biopsies of normal colon and Crohn's disease were evaluated. We examined 77 colonic mucosa specimens taken from patients undergoing elective resection for benign and malignant colonic disease. uPA and PAI-1 were measured using a monoclonal antibody-based ELISA kit (American Diagnostica, Greenwich, CT) and expressed as ng/mg extract protein. Intra- and interassay controls of uPA gave CV = 3-4% and CV = 8-9%, respectively, while those for PAI-1 were 6-7% and 10-11%, respectively. The Mann-Whitney test showed that both uPA and PAI-1 expression were significantly higher in colon cancer, chronic ulcerative colitis, and Crohn's disease than in normal colon. uPA in familial polyposis samples was similar to that of normal colon, while PAI-1 was much lower than in normal colon. Neither patient age nor sex appeared to influence the expression of these potential markers in any tissue. The pattern of uPA and PAI-1 expression in normal, benign and malignant colon suggests these proteins deserve further consideration as markers for assessing colon carcinoma risk.

Adenomatous Polyposis Coli↗

Protein kinase C isoforms in human and rat colonic mucosa.

The protein kinase C (PKC) family of enzymes plays a key role in the regulation of cellular events, including cell proliferation and differentiation. Work from our laboratory has shown that the effects of dietary fat and fiber on colonic cell proliferation were positively correlated with membrane/cytosol PKC activity ratios (Chapkin et al., 1993, J. Nutr. 123, 649-655). The presence and subcellular distribution of specific PKC isoforms in rat and human colon were therefore determined in cytosolic and membrane extracts. Tissue extracts were probed with antibodies to individual PKC isoforms. PKC alpha, beta, delta, epsilon, and zeta were detected in both rat and human colonic mucosa, while PKC eta was detected in human colonic mucosa only. PKC alpha, beta, and zeta were predominantly localized in the cytosolic fraction, whereas the majority of PKC delta, epsilon, and eta were found in the membrane-associated fraction. Presence of mRNA for individual PKC isoforms was determined by reverse transcriptase PCR (RT-PCR). Using rat colonic mucosa, mRNA for PKC alpha, beta, delta, epsilon, eta, and zeta were detected by RT-PCR with identity confirmed by sequencing. The relative steady-state levels of PKC isoforms in human colon adenocarcinoma as compared with normal colonic mucosa were determined, with adenocarcinomas having higher amounts of cytosolic PKC beta, delta, epsilon, eta, and zeta. PKC isoforms were also detected in viable, exfoliated colonic cells isolated from human feces, demonstrating that this noninvasive method can be utilized to examine PKC expression in colonic cells. These results demonstrate that colonic mucosa expresses both calcium-dependent (classical) and calcium-independent (novel and atypical) PKC isoforms with distinct subcellular distributions for each. The dynamics of these PKC isoforms may have implications in the development of colon carcinogenesis.

Adenocarcinoma↗

Colonic epithelium reactive monoclonal antibodies. Identification and immunohistochemical localization of the target epitopes.

We have produced a small library of colonic mucosa and colorectal carcinoma reactive monoclonal antibodies (MoAbs) by immunizations with extracts of human colon cancer tissue and a human colon cancer cell line. Hybridoma supernatants were tested on (normal and neoplastic) human tissues by immunoperoxidase methods to evaluate organ or tissue specificity. Initial biochemical characterization of the target antigens was performed by gelpermeation chromatography, Western blotting and competition assays. Based upon the immunoreactivity patterns and the characteristics of the antigen four groups of MoAbs could be distinguished. The first group concerns the antibodies PARLAM 3, 9 and 10. These antibodies react with an 87 kDa protein moiety in high molecular weight (2-5 x 10(6) Da) glycoproteins. In intestinal and colon mucosa these antibodies showed diffuse binding with goblet cells. In colon carcinoma decreased reactivity with these MoAbs was found. The second group consists of antibodies PARLAM 8, 12 and 13. These antibodies react with large (greater than 5 x 10(6) Da) glycoproteins, most likely with carbohydrate epitopes. By immunohistochemistry in normal colon mucosa the antibodies all show granular supranuclear reactivity with goblet cells. These antibodies show increased reactivity with colon adenomas and adenocarcinomas. A third group is formed by PARLAM 2, which also reacts with a large (greater than 5 x 10(6) Da) glycoprotein, showing a granular distribution in goblet cells. In colon carcinomas more extensive expression is found than in normal colonic mucosa. Finally, the fourth group consists of PARLAM 11, which also reacts with a large (greater than 5 x 10(6) Da) glycoprotein, located in the brush border of colonic columnar cells. These antibodies might be useful tools for the analysis of the expression of mucin related glycoproteins in normal, preneoplastic and neoplastic colon mucosa.

Adenocarcinoma↗