Analysis of stromal signatures in the tumor microenvironment of ductal carcinoma in situ.

Recent advances in the study of the tumor microenvironment have revealed significant interaction between tumor cells and their surrounding stroma in model systems. We have previously shown that two distinct stromal signatures derived from a macrophage (CSF1) response and a fibroblastic (DTF-like) response are present in subsets of invasive breast cancers and show a correlation with clinical outcome. In the present study we explore whether these signatures also exist in the stroma of ductal carcinoma in situ (DCIS). We studied the signatures by both gene expression profile analysis of a publically available data set of DCIS and by immunohistochemistry (IHC) on a tissue microarray of DCIS and invasive breast cancer cases. Both the gene expression and immunohistochemical data show that the macrophage response and fibroblast expression signatures are present in the stroma of subsets of DCIS cases. The incidence of the stromal signatures in DCIS is similar to the incidence in invasive breast cancer that we have previously reported. We also find that the macrophage response signature is associated with higher grade DCIS and cases which are ER and PR negative, whereas the fibroblast signature was not associated with any clinicopathologic features in DCIS. A comparison of 115 matched cases of DCIS and invasive breast cancer found a correlation between the type of stromal response in DCIS and invasive ductal carcinoma (IDC) within the same patient for both the macrophage response and the fibroblast stromal signatures (P = 0.03 and 0.08, respectively). This study is a first characterization of these signatures in DCIS. These signatures have significant clinicopathologic associations and tend to be conserved as the tumor progresses from DCIS to invasive breast cancer.

Variable hearing impairment in a DFNB2 family with a novel MYO7A missense mutation.

Myosin VIIA mutations have been associated with non-syndromic hearing loss (DFNB2; DFNA11) and Usher syndrome type 1B (USH1B). We report clinical and genetic analyses of a consanguineous Iranian family segregating autosomal recessive non-syndromic hearing loss (ARNSHL). The hearing impairment was mapped to the DFNB2 locus using Affymetrix 50K GeneChips; direct sequencing of the MYO7A gene was completed. The Iranian family (L-1419) was shown to segregate a novel homozygous missense mutation (c.1184G>A) that results in a p.R395H amino acid substitution in the motor domain of the myosin VIIA protein. As one affected family member had significantly less severe hearing loss, we used a candidate approach to search for a genetic modifier. This novel MYO7A mutation is the first reported to cause DFNB2 in the Iranian population and this DFNB2 family is the first to be associated with a potential modifier. The absence of vestibular and retinal defects, and less severe low frequency hearing loss, is consistent with the phenotype of a recently reported Pakistani DFNB2 family. Thus, we conclude this family has non-syndromic hearing loss (DFNB2) rather than USH1B, providing further evidence that these two diseases represent discrete disorders.

Building a clinical cardiothoracic surgical program: a multi-institutional model.

Building a multi-institutional cardiothoracic surgical program has the same guiding principles and values as a traditional single institutional program: ensuring high-quality patient care, training and fostering residents, recruiting and retaining quality faculty, and contributing to basic and clinical research. With a well-designed infrastructure and support system, this more complicated type of organization may permit academic cardiothoracic surgical programs to compete effectively and grow in a constantly changing economic and political environment.

Ubiquitous presence of a mecA homologue in natural isolates of Staphylococcus sciuri.

In an effort to explore the origin and/or reservoirs of the genetic determinant(s) of methicillin resistance in Staphylococcus aureus, we examined over 200 strains representing 13 different species within the genus Staphylococcus for the presence of the mecA gene, using a DNA probe internal to this gene prepared from a methicillin-resistant strain of S. aureus. Occasional mecA- positive isolates were detected among several staphylococcal species. On the other hand, each one of the 134 isolates of Staphylococcus sciuri, a species considered taxonomically the most primitive among staphylococci and found primarily on rodents and primitive mammals, gave positive reaction with the DNA probe when tested under conditions of high stringency. About two thirds (99) of these isolates, all of which belonged to S. sciuri subspecies "sciuri," as well as 9 of the 11 species carnaticum isolates, showed only marginal, if any, resistance to methicillin (minimal inhibitory concentration of 0.75-6.0 micrograms/ml), while most of the remaining isolates that belonged to the subspecies "rodentius" (13 isolates in all) expressed antibiotic resistance with a heterogeneous phenotype similar to those seen in many methicillin-resistance strains of S. aureus In SmaI digests of chromosomal DNA isolated from such "methicillin-resistant S. aureus-like" strains, the mecA probe hybridized with DNA fragments in the range of 145-180 kb, while in subspecies "sciuri" and carnaticum isolates the mecA hybridizing fragment was located in the SmaI fragment with the highest molecular size (> or = 400 kb). A DNA probe comprising an internal sequence to the regulatory gene mecI from Staphylococcus epidermidis identified the presence of sequences with low degree of homology in isolates of the three S. sciuri subspecies. The mecA-reacting sequences in these bacteria differed from mecA of S. aureus in several respects (e.g., by the absence of a ClaI restriction site from mecA of subspecies "sciuri" and carnaticum, and in some isolates of subspecies "rodentius." The uniform presence of mecA in each one of a large number of S. sciuri strains belonging to distinct ribotypes and macrorestriction patterns and recovered over a 20-year period from a wide variety of animal sources and geographic sites suggests that mecA may be a native genetic element with an as yet unidentified physiologic function in this staphylococcal species.

WHO-sponsored international collaborative study to evaluate methods for subtyping Listeria monocytogenes: restriction fragment length polymorphism (RFLP) analysis using ribotyping and Southern hybridization with two probes derived from L. monocytogenes chromosome.

Seven laboratories participated in a WHO-sponsored international collaborative study, to evaluate methods for subtyping Listeria monocytogenes, by performing restriction fragment length polymorph sm (RFLP) analysis-based subtyping of an international study set of 80 strains of L. monocytogenes that included 22 epidemiologically related groups. The RFLP analysis was done by Southern hybridization with one of two types of probes found in multiple copies on the chromosome of L. monocytogenes. Six laboratories performed ribotyping. These laboratories used EcoRI enzyme to restrict the L. monocytogenes DNA and ribosomal RNA or DNA as the probe for Southern hybridizations. The seventh laboratory used Ncil to restrict the DNA, and two probes, one randomly cloned and the other containing repeat sequences cloned from L. monocytogenes DNA. The overall discriminating power of ribotyping, as estimated by calculation of Simpson's index of diversity, ranged from 0.83 to 0.88 for the six laboratories. The discriminating power of the combination of two probes used by Laboratory 7 was 0.91. Ribotyping and the cloned probes used by Laboratory 7 discriminated poorly between serotype 4b strains. Neither method identified three atypical strains (identified by other subtyping methods) included in three apparently epidemiologically related groups. Ribotyping did not discriminate between strains of serotypes 4b and 4b(X) in one epidemiologically related group of strains; one cloned probe used by Laboratory 7 discriminated between these strains. Intra-laboratory reproducibilities for the seven laboratories ranged from 80.0 to 100%. as determined by their abilities to correctly identify 11 pairs of duplicate strains included in the study set. Inter-laboratory reproducibilities were generally very good considering that no attempt was made to standardize protocols used by the participants.

Plant resistance components of two greenbug (Homoptera: Aphididae) resistant wheats.

Several biotypes of the greenbug, Schizaphis graminum (Rondani), attack winter wheat, Triticum aestivum L., on the Southern Plains every year. Two wheat germplasm sources of resistance ('Largo' and 'GRS 1201') have been developed that provide protection against the three predominant greenbug biotypes (E, I, and K). Each source has agronomic and end-use quality advantages and disadvantages for the breeder to consider in choosing a greenbug-resistant breeding line. We compared these two germplasms to determine their levels of resistance against biotype E. Components of resistance (i.e., antibiosis, antixenosis, and tolerance) were measured on seedlings of GRS 1201, Largo, and 'TAM W-101' (a susceptible control). Several aphid and plant measurements (e.g., total number of aphids produced per plant, aphid selection preferences, and plant damage ratings) were recorded for each plant entry. Select data recorded for each resistance component were normalized and combined to derive a plant resistance index for each wheat entry. Results indicated that GRS 1201 had a higher level of combined resistance components than did Largo, followed by TAM W-101, the susceptible control. These data provide additional information for the breeder to consider in selecting a greenbug-resistant breeding line.

Efficacy of pyramiding greenbug (Homoptera: Aphididae) resistance genes in wheat.

Durable resistance to greenbug, Schizaphis graminum (Rondani), in wheat is a goal of wheat improvement teams, and one that has been complicated by the regular occurrence of damaging biotypes. Simulation modeling studies suggest that pyramiding resistance genes, i.e., combining more than one resistance gene in a single cultivar or hybrid, may provide more durable resistance than sequential releases of single genes. We examined this theory by pyramiding resistance genes in wheat and testing a series of greenbug biotypes. Resistance genes Gb2, Gb3, and Gb6, and pyramided genes Gb2/Gb3, Gb2/Gb6, and Gb3/Gb6 were tested for effectiveness against biotypes E, F, G, H, and I. By comparing reactions of plants with pyramided genes to those with single resistance genes, we found that pyramiding provided no additional protection over that conferred by the single resistance genes. Based on the results of this test, we concluded that the sequential release of single resistance genes, combined with careful monitoring of greenbug population biotypes, is the most effective gene deployment strategy for greenbug resistance in wheat.

Policy for controlling pain after surgery: effect of sequential changes in management.

OBJECTIVE: To observe the effects of introducing an acute pain service to the general surgical wards of a large teaching hospital. DESIGN: A study in seven stages: (1) an audit of current hospital practice succeeded by the sequential introduction to the general surgical wards of (2) pain assessment charts; (3) an algorithm to allow more frequent use of intramuscular analgesia; (4) increased use of local anaesthetic techniques of wound infiltration and nerve blocks; (5) an information sheet for patients about postoperative pain; (6) the introduction of patient controlled analgesia; (7) a repeat audit of hospital practice. Data were collected on each patient 24 hours after operation. SETTING: University Hospital of Wales, which has both district general and tertiary referral functions. PATIENTS: 2035 patients over nine months from all surgical specialties (excluding cardiac) at the hospital. General surgical operations were studied in detail and separated into major, intermediate, and minor for data collection. MAIN OUTCOME MEASURES: A change in the median visual analogue pain scores 24 hours after surgery for pain during relaxation, pain on movement, and pain on deep inspiration at each stage of the study. RESULTS: There was a reduction in median visual analogue scores during the study. The median (95% confidence interval) scores for pain during relaxation decreased from 45 (34 to 53) in stage 1 to 16 (10 to 20) in stage 7 for major surgical procedures. Pain on movement decreased from 78 (66 to 80) to 46 (38 to 48), and pain on deep inspiration decreased from 64 (48 to 78) to 36 (31 to 38). The reductions in median scores for intermediate and minor operative procedures showed similar patterns. CONCLUSIONS: The introduction of an acute pain service to the general surgical wards led to considerable improvement in the level of postoperative pain as assessed by visual analogue scores. Simple techniques of regular pain assessment and the more frequent use of intramuscular analgesia as a result of using an algorithm were particularly effective.

Deep Vein Thrombosis in Marathon Runners.

In brief: In these case reports of two highly trained runners, deep vein thrombosis (DVT) was initially diagnosed as an intramuscular hematoma and a Baker's cyst. Both runners responded well to anticoagulant therapy and were able to resume their training programs. Even though otherwise healthy, conditioned patients are not typical candidates, the authors urge that DVT be considered in the differential diagnosis of athletes with leg pain or swelling because of the threat of pulmonary embolism.

Rapid identification of bacteria on the basis of polymerase chain reaction-amplified ribosomal DNA spacer polymorphisms.

To facilitate genus and species level identification of a broad range of bacteria without the requirement of presumptive identification, we have developed a unified set of primers and polymerase chain reaction conditions to amplify spacer regions between the 16S and 23S genes in the prokaryotic rRNA genetic loci. Spacer regions within these loci show a significant level of length and sequence polymorphism across both genus and species lines. A generic pair of priming sequences was selected for the amplification of these polymorphisms from highly conserved sequences in the 16S and 23S genes occurring adjacent to these polymorphic regions. This single set of primers and reaction conditions was used for the amplification of 16S-23S spacer regions for over 300 strains of bacteria belonging to eight genera and 28 species or serotypes, including Listeria, Staphylococcus, and Salmonella species and additional species related to these pathogenic organisms. When the spacer amplification products were resolved by electrophoresis, the resulting patterns could be used to distinguish all of the species of bacteria within the test group. Unique elements in the amplification product patterns generally clustered at the species level, although some genus-specific characteristics were also observed. On the basis of the results obtained with our test group of 300 bacterial strains, amplification of the 16S-23S ribosomal spacer region is a suitable process for generating a data base for use in a polymerase chain reaction-based identification method, which can be comprehensively applied to the bacterial kingdom.

Generation of clonal diversity by sexual reproduction in the greenbug, Schizaphis graminum.

In the United States, the greenbug, Schizaphis graminum (Rondani), reproduces primarily by apomictic parthenogenesis. Although a periodic sexual cycle exists, the extent to which it occurs naturally and its influence on the genetic variability of greenbug populations is unclear. Length variation in the intergenic spacer (IGS) of the rRNA cistron in the greenbug indicates that populations are made up of many genetically distinct clones. Previous laboratory studies have shown the stability of the IGS within parthenogenetic clones. By inducing the sexual reproductive cycle of the greenbug, we conducted both Intra- and inter-clone matings and studied the inheritance of the IGS in the offspring. In both mating schemes, rearrangements in the IGS were apparent. IGS diversity found among the offspring could be attributed to unequal cross-over and probably other molecular drive events during meiosis. Periodic sexual reproduction is a primary mechanism for the generation and maintenance of genetic variability in greenbug populations, and explains the level of clonal diversity found in previous studies.

In vitro release of amoxycillin from lipophilic suppositories.

The in vitro release characteristics of amoxycillin from different lipophilic suppository bases were investigated using the USP rotating basket method. Suppositories containing 250 mg amoxycillin were prepared in theobroma oil and in the semi-synthetic bases Witepsol W35, Suppocire A32, Novata BD, and Novata 299. Both freshly prepared and 1-month-old suppositories were tested. Analysis of amoxycillin was performed using a validated high-performance liquid chromatographic (HPLC) technique. Release profiles differed significantly between bases, with the greatest amount of amoxycillin being released from both newly made and 1-month-old Novata BD bases (87.57 +/- 8.18 and 99.66 +/- 6.63%, respectively), and the lowest amount released from the newly manufactured theobroma suppositories (8.82 +/- 0.75%) and the 1-month-old Suppocire A32 suppositories (7.78 +/- 0.27%).

Suprofen compared to dextropropoxyphene hydrochloride and paracetamol (Cosalgesic) after extraction of wisdom teeth under general anaesthesia.

In a randomised double-blind trial in postoperative ambulant day case dental patients suprofen 200 mg (29 patients) was compared with dextropropoxyphene hydrochloride 65 mg and paracetamol 650 mg (Cosalgesic, 28 patients) both available four times daily for 3 days. Suprofen was better than cosalgesic in the patients' opinion of initial (p = 0.01) and overall pain relief (p = 0.08) compared to Cosalgesic and the second night's sleep was better (p = 0.01). Side effects were reported in six suprofen patients and 10 cosalgesic patients (two suffering from vomiting withdrew). Suprofen, a non-steroidal anti-inflammatory drug is as good as, or better than, a widely used opioid-paracetamol mixture for ambulant patients with postoperative dental pain.