Fewer than 12 lymph nodes can be expected in a surgical specimen after high-dose chemoradiation therapy for rectal cancer.

PURPOSE: Lymph node harvest of >or=12 has been adopted as a marker for adequacy of resection for colorectal cancer. We have noted a paucity of lymph nodes in rectal cancer specimens after neoadjuvant therapy, positing that the number of lymph nodes depends on the response to radiation and may not be an appropriate benchmark. Our purpose was to determine whether the number of lymph nodes harvested after neoadjuvant therapy is a useful quality indicator. METHODS: A database of rectal cancer patients was queried to identify patients undergoing total mesorectal excision after neoadjuvant chemoradiation between January 1997 and August 2007. We compared patients with <12 lymph nodes to those with >or=12 lymph nodes relative to multiple patient and treatment factors. RESULTS: One hundred seventy-six patients were identified (119 men; mean age, 60.4 y (range, 22-87)). Mean lymph node harvest was 10.1 (range, 1-38). Only 28% had >or=12 lymph nodes and 32% had <6 lymph nodes. There was no statistically significant difference in lymph node harvest relative to radiation dosage, age, tumor response, or type of surgery. There was no correlation between the number of lymph nodes harvested and the number of nodes positive for cancer. CONCLUSIONS: With a standardized surgical technique and pathologic evaluation, the number of lymph nodes present after neoadjuvant chemoradiation and total mesorectal excision for rectal cancer varies greatly.

Evidence that hereditary pancreatitis is genetically heterogeneous disorder.

Hereditary pancreatitis (HP) is an autosomal dominant disorder characterized by recurrent acute attacks of severe abdominal pain with an onset in early childhood. Many HP patients progress to complicated chronic pancreatitis and/or pancreatic cancer. Initially, a single mutation R117H in the cationic trypsinogen gene was detected in all affected members of five unrelated HP families. Further studies identified a second mutation (N21L) in two HP families without the R117H mutation. Before the association between cationic trypsinogen and HP was found, we detected a cystic fibrosis transmembrane conductance regulator (CFTR) gene mutation (L327R) in all affected individuals of a family with HP. We therefore performed a mutational analysis for R117H and N21L in cationic trypsinogen in this and three additional unrelated families with HP. The R117H mutation was detected in all 9 affected members of three HP families and in 3 asymptomatic but at-risk relatives. However, neither the R117H nor the N21L mutation in the cationic trypsinogen were found in the HP family with the L327R alteration in CFTR. The L327R allele segregates with the disease within this HP family and was not detected on 360 unrelated Caucasian non-CF chromosomes. Although close to 800 different mutations have been detected in the CF gene of cystic fibrosis patients, L327R is a new alteration, not yet reported in connection with CF. The results of this study indicate that the CFTR gene may play a role in the etiology of minority of cases with HP and suggest that hereditary pancreatitis is genetically heterogeneous disease.

Molecular basis of cystic fibrosis in the Republic of Macedonia.

Eighty-three cystic fibrosis (CF) patients and their families, belonging to various ethnic groups living in the Republic of Macedonia were studied for molecular defects in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, and for the associated extragenic marker loci XV-2c and KM19. The DNA methodology used included characterization of CFTR mutations in 19 exons (and flanking sequences) of the gene and analysis of distribution of the XV-2c/KM19 haplotypes among normal (N) and CF chromosomes by polymerase chain reaction (PCR) amplification followed by dot blot hybridization, restriction digestion, single-strand conformational polymorphism, constant denaturing gel electrophoresis, denaturing gradient gel electrophoresis, and sequencing. We identified 58.4% (97/166) of the CF chromosomes. Nine different CFTR gene mutations, including three novel ones, were found. Eight known and one new CFTR intragene polymorphisms were also characterized. The haplotype analysis of the XV-2c/TaqI and KM19/PstI polymorphic loci have shown that haplotype C is the most frequently found haplotype among the non-deltaF508 CF chromosomes from Macedonia (36.5%). The results demonstrate the broad heterogeneity of CF origin in this part of the Balkan Peninsula.

Cystic fibrosis gene mutations detected in hereditary pancreatitis.

To determine the potential role of the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) gene in the etiology of Hereditary Pancreatits (HP), we analyzed this gene in two HP families. PCR-SSCP analysis was employed, followed by direct sequencing of DNA samples showing alterations by SSCP. A new alteration not yet detected in connection with CF was detected in one HP family. The alteration is in exon 7 and causes an amino acid change from Leu to Arg at position 327 (L327R). L327R segregates with the disease within the family. We discovered another novel alteration (V1190P) in an HP patient of the second family. These results indicate that CFTR gene mutations may be involved in the etiology of HP and represent a challenge for further study of the role of (CTFR) in other digestive diseases.

Screening for CF mutations in adult cystic fibrosis patients with a directed and optimized SSCP strategy.

Twenty adolescent and adult cystic fibrosis (CF) patients have been studied for the presence of mutations in the CFTR gene. Mutations other than deltaF508 have been detected by comparison to the single-stranded conformation polymorphism (SSCP) pattern of known mutations in eight exons, in which 80% of the more common mutations are present. Each mutation was confirmed by direct sequencing. For each of the analyzed exons, optimal SSCP conditions have been determined that allow all available known mutations in that exon to be distinguished from each other. This approach allowed mutations to be defined in 75% of the non deltaF508 alleles and 92% of all CF alleles in this cohort.

Expression of the cystic fibrosis transmembrane conductance regulator gene in the respiratory tract of normal individuals and individuals with cystic fibrosis.

The most common mutation of the cystic fibrosis transmembrane conductance regulator gene, CFTR, associated with the clinical disorder cystic fibrosis (CF) is called "delta Phe508," a triple-base deletion resulting in loss of phenylalanine at residue 508 of the predicted 1480-amino acid CFTR protein. In the context that the lung is the major site of morbidity and mortality in CF, we evaluated airway epithelial cells for CFTR mRNA transcripts in normal individuals, normal-delta Phe508 heterozygotes, and delta Phe508 homozygotes to determine if the normal and delta Phe508 CFTR alleles are expressed in the respiratory epithelium, to what extent they are expressed, and whether there are relative differences in the expression of the normal and abnormal alleles at the mRNA level. Respiratory tract epithelial cells recovered by fiberoptic bronchoscopy with a cytology brush demonstrated CFTR mRNA transcripts with sequences appropriately reflecting the normal and delta Phe508 CFTR alleles of the various study groups. CFTR gene expression quantified by limited polymerase chain reaction amplification showed that in normal individuals, CFTR mRNA transcripts are expressed in nasal, tracheal, and bronchial epithelial cells at approximately 1-2 copies per cell, more than 100-fold greater than in pharyngeal epithelium. Importantly, allele-specific hybridization studies demonstrated that the normal and delta Phe508 CFTR alleles are expressed in the respiratory epithelium in similar amounts.

Aerosol alpha 1-antitrypsin treatment for cystic fibrosis.

In cystic fibrosis neutrophil-dominated inflammation on the respiratory epithelial surface results in a chronic epithelial burden of the destructive enzyme, neutrophil elastase. alpha 1-antitrypsin (alpha 1AT), the main inhibitor of neutrophil elastase in lung, was given in aerosol form to 12 cystic fibrosis patients. It suppressed neutrophil elastase in the respiratory epithelial lining fluid (ELF) and restored the ELF anti-neutrophil elastase capacity when ELF alpha 1AT reached 8 mumol/l. This treatment also reversed the inhibitory effect of cystic fibrosis ELF on pseudomonas killing by neutrophils, which suggests that it may augment host defence in cystic fibrosis.

Isolation and characterization of mucin from the serum of cystic fibrosis patients.

Using monoclonal antibody 19-9, elevated levels of the sialyl Lea antigen (NeuAc alpha 2-3Gal beta 1-3[Fuc alpha 1-4]GlcNac beta 1-3R) are detected in serum from most cystic fibrosis patients. We now report further characterization of the serum antigen and evidence that it is on a mucin glycoprotein and not on glycolipids. The antigen has an apparent molecular weight greater than 2 X 10(6) by gel filtration on Sephacryl S-400. On density gradient centrifugation, the antigen has a density of 1.54 g/ml in cesium chloride and 1.42 g/ml in cesium chloride/4 M guanidine HCl. Immunostaining with monoclonal antibody 19-9 of lipid extracts from cystic fibrosis patient serum and erythrocytes does not detect any antigen on glycolipids. The antigen was purified by gel filtration and density gradient centrifugation. After tritium labeling of the sialic acid residues, sodium dodecyl sulfate gel electrophoresis separates two subunits with apparent molecular weights of 200,000 and 400,000. All of the labeled sialic acid is released as low molecular weight oligosaccharides after mild alkaline borohydride degradation. The purified antigen contains fucose, galactosamine, glucosamine, and galactose but no mannose and is enriched in the amino acids threonine, serine, glycine, proline, and alanine. The purified antigen binds several antibodies recognizing epitopes common to many mucins. Thus, the physical, biochemical, and immunochemical properties of the purified antigen indicate that the sialyl Lea antigen is present on mucins in the serum of cystic fibrosis patients.

Serum mucin-associated antigen levels of cystic fibrosis patients are related to their ages and clinical statuses.

Mucin levels are generally elevated in sera from many cystic fibrosis (CF) patients as measured by radioimmunoassay using monoclonal antibody 19-9, which is directed against the mucin-associated sialyl Lea antigen. Antibody 19-9 can only be used to measure mucin-associated antigen levels in those patients who are genetically able to make detectable levels of mucin-associated sialyl Lea epitope. Serial studies of 20 patients followed over 3-5 y showed that their serum mucin-associated antigen levels varied directly with respect to the severity of their disease and inversely with their Shwachman-Kulczycki clinical scores (p less than 0.001) and Brasfield chest roentgenographic scores (p less than 0.02). Serum mucin-associated antigen levels in samples from 89 CF patients were generally higher in the older patients (p less than 0.025). Serum mucin-associated antigen levels of CF patients who were colonized with Pseudomonas aeruginosa did not significantly differ from those of uninfected CF patients. The mean serum mucin-associated antigen level of CF patients colonized with Pseudomonas was higher than the mean mucin level of six non-CF bronchiectatic patients whose lungs were colonized with Pseudomonas (p = 0.053). Serum mucin-associated antigen levels are thus related to CF patients' ages and clinical statuses.

A serum test for cystic fibrosis using monoclonal antibody 19-9.

Monoclonal antibody 19-9 detects a sialosylated Lea antigen with the following sugar sequence: NeuNAc alpha 2-3Gal beta 1-3[Fuc alpha 1-4]GlcNAc beta 1-3Gal. . . . This antigen is detected as a mucin in the sera of many patients with gastrointestinal and pancreatic cancer. Elevated levels of sialosylated Lea antigen are also detected in serum from 14 of 16 patients with cystic fibrosis (87%). One of the two negative patients belongs to the Le(a-b-) blood group and so is unable to synthesize the sialosylated Lea antigen. The high percentage of cystic fibrosis patients with elevated sialosylated Lea antigen suggests that the 19-9 antibody may be useful for diagnosis of cystic fibrosis. Antibodies to other sialosylated carbohydrates in mucins may also be useful for detection of cystic fibrosis and may allow diagnosis of patients belonging to the Le(a-b-) blood group.

Appendiceal abscess in cystic fibrosis. A diagnostic challenge.

A patient with cystic fibrosis who developed appendicitis, rupture, and a periappendiceal abscess is presented. A retrospective chart review revealed 5 other cases that demonstrate a spectrum of clinical presentation of periappendiceal abscess in patients with cystic fibrosis. Three patients were symptomatic for less than 5 days, but the remaining 3 patients were symptomatic for 8, 12, and 30 days before diagnosis. There were two deaths due to respiratory failure. Other complications included a perirectal fistula and 2 cases of recurrent abscess. This demonstrates the difficulty with which this diagnosis is reached in this patient population and the relatively high incidence of abscess formation compared with normal populations. A retrospective autopsy review of 51 cystic fibrosis patients showed that in 49 of 51 instances, the mucosa of the appendix was hyperplastic, and the mucosal glands were distended with eosinophilic secretions. In 12 cases (24%), the appendix itself was grossly firm, dilated, and distended, although the mucosal wall was free of inflammation. This lends credence to the suggestion that these inspissated secretions may be protective against the occurrence of appendicitis, the incidence of which may be as low as 1%-2% among cystic fibrosis patients.

Scoliosis in cystic fibrosis--an appraisal.

An unusually high prevalence (10%) of scoliosis is described in a series of 151 patients aged four years and older with cystic fibrosis. The scolioses were of the late onset (juvenile and adolescent) type, being typically thoracic with the curve convex to the right, although there was no significant preference for either sex. No direct relationship was found between the spinal curvature and the severity or distribution of the lung disease, although the worse scolioses tended to occur in patients with relatively severe pulmonary involvement. There was no evidence of metabolic bone disease as a predisposing cause. Some indication of a familial tendency towards scoliosis was apparent, and a genetic or constitutional basis is postulated with an unknown precipitating factor.