Comprehensive epigenomic profiling of human alveolar epithelial differentiation identifies key epigenetic states and transcription factor co-regulatory networks for maintenance of distal lung identity.

BACKGROUND: Disruption of alveolar epithelial cell (AEC) differentiation is implicated in distal lung diseases such as chronic obstructive pulmonary disease, idiopathic pulmonary fibrosis, and lung adenocarcinoma that impact morbidity and mortality worldwide. Elucidating underlying disease pathogenesis requires a mechanistic molecular understanding of AEC differentiation. Previous studies have focused on changes of individual transcription factors, and to date no study has comprehensively characterized the dynamic, global epigenomic alterations that facilitate this critical differentiation process in humans. RESULTS: We comprehensively profiled the epigenomic states of human AECs during type 2 to type 1-like cell differentiation, including the methylome and chromatin functional domains, and integrated this with transcriptome-wide RNA expression data. Enhancer regions were drastically altered during AEC differentiation. Transcription factor binding analysis within enhancer regions revealed diverse interactive networks with enrichment for many transcription factors, including NKX2-1 and FOXA family members, as well as transcription factors with less well characterized roles in AEC differentiation, such as members of the MEF2, TEAD, and AP1 families. Additionally, associations among transcription factors changed during differentiation, implicating a complex network of heterotrimeric complex switching in driving differentiation. Integration of AEC enhancer states with the catalog of enhancer elements in the Roadmap Epigenomics Mapping Consortium and Encyclopedia of DNA Elements (ENCODE) revealed that AECs have similar epigenomic structures to other profiled epithelial cell types, including human mammary epithelial cells (HMECs), with NKX2-1 serving as a distinguishing feature of distal lung differentiation. CONCLUSIONS: Enhancer regions are hotspots of epigenomic alteration that regulate AEC differentiation. Furthermore, the differentiation process is regulated by dynamic networks of transcription factors acting in concert, rather than individually. These findings provide a roadmap for understanding the relationship between disruption of the epigenetic state during AEC differentiation and development of lung diseases that may be therapeutically amenable.

Primary colorectal linitis plastica presenting as rapid acute deterioration: a diagnostic dilemma.

We present a rare case of primary colorectal linitis plastica presenting as an acute admission to hospital with a wide range of systemic symptoms, sudden rapid deterioration and subsequent mortality. A postmortem examination revealed a primary linitis plastica of the colon and rectum with diffuse metastatic disease. To our knowledge, this is the first report of primary colorectal linitis plastica presenting as an acute deterioration as a result of extensive metastatic disease.

Improving diagnostic accuracy in clinically ambiguous paediatric appendicitis: a retrospective review of ultrasound and pathology findings with focus on the non-visualised appendix.

OBJECTIVE: To compare pre-operative ultrasound to histopathological results and retrospectively assess the diagnostic accuracy of ultrasound in paediatric appendicitis. METHODS: 5 year review of all appendectomies performed in patients <16 years old in a tertiary referral university hospital. 983 patients had an appendicectomy over the time period while 189 patients had a preoperative ultrasound. We retrospectively reviewed all of the preoperative imaging in conjunction with the reports for the 189 patients; our aim was to determine the sensitivity of preoperative ultrasound for the diagnosis of acute appendicitis. RESULTS: Of the 189 patients who had an ultrasound, 102 had histology positive for appendicitis and 87 had normal histology. Sensitivity overall was 72.55% [95% confidence interval (CI) 62.82 to 80.92] and specificity was 77.01% (95% CI 66.75 to 85.36). A suggested ultrasound diagnosis of appendicitis made positive pathology three times more likely and a normal ultrasound made positive pathology three times less likely [positive-predictive value 3.16 (95% CI 2.11 to 4.72) negative predictive value 0.36 (95% CI 0.25 to 0.50)]. 77% (67/87) of the patients whose pathology was ultimately normal had an ultrasound which was either normal or suggested an alternative diagnosis. However, in the 33 (17%) of patients with a non-visualised appendix, no secondary signs of inflammation or alternative diagnosis 16 (48%) had pathologically confirmed appendicitis. CONCLUSION: Ultrasound has the potential to improve diagnostic accuracy in clinically ambiguous appendicitis. ADVANCES IN KNOWLEDGE: This paper furthers the evidence on the efficacy of ultrasound as a diagnostic tool in acute appendicitis in children, especially when the diagnosis is clinically equivocal. It also sheds further light on the "non-visualized appendix" with almost half of these patients having pathologically confirmed appendicitis; meaning advanced imaging with CT or MR may be indicated in this cohort.

Magnetic Resonance Image-Guided Radiotherapy (MRIgRT): A 4.5-Year Clinical Experience.

AIMS: Magnetic resonance image-guided radiotherapy (MRIgRT) has been clinically implemented since 2014. This technology offers improved soft-tissue visualisation, daily imaging, and intra-fraction real-time imaging without added radiation exposure, and the opportunity for adaptive radiotherapy (ART) to adjust for anatomical changes. Here we share the longest single-institution experience with MRIgRT, focusing on trends and changes in use over the past 4.5 years. MATERIALS AND METHODS: We analysed clinical information, including patient demographics, treatment dates, disease sites, dose/fractionation, and clinical trial enrolment for all patients treated at our institution using MRIgRT on a commercially available, integrated 0.35 T MRI, tri-cobalt-60 device from 2014 to 2018. For each patient, factors including disease site, clinical rationale for MRIgRT use, use of ART, and proportion of fractions adapted were summated and compared between individual years of use (2014-2018) to identify shifts in institutional practice patterns. RESULTS: Six hundred and forty-two patients were treated with 666 unique treatment courses using MRIgRT at our institution between 2014 and 2018. Breast cancer was the most common disease, with use of cine MRI gating being a particularly important indication, followed by abdominal sites, where the need for cine gating and use of ART drove MRIgRT use. One hundred and ninety patients were treated using ART in 1550 fractions, 67.6% (1050) of which were adapted. ART was primarily used in cancers of the abdomen. Over time, breast and gastrointestinal cancers became increasingly dominant for MRIgRT use, hypofractionated treatment courses became more popular, and gastrointestinal cancers became the principal focus of ART. DISCUSSION: MRIgRT is widely applicable within the field of radiation oncology and new clinical uses continue to emerge. At our institution to date, applications such as ART for gastrointestinal cancers and accelerated partial breast irradiation (APBI) for breast cancer have become dominant indications, although this is likely to continue to evolve.

Dependence and psychosis with 4-methylmethcathinone (mephedrone) use.

'Legal highs' are recreational drugs sold over the internet and the so-called 'head shops' all over the UK. They are freely available to buy and use as they are not covered by the Misuse of Drugs Act 1971. Mephedrone (4-methylmethcathinone) was sold as a 'legal high' until 17 April 2010 when it was made a class B drug under the Misuse of Drugs Act 1971. Numerous deaths and self-harm has been associated with mephedrone use. Effects of mephedrone are reported to be empathogenic similar to 3,4-methylenedioxymethamphetamine (MDMA, ecstasy) and stimulant properties similar to cocaine. Not much is known of the effects of mephedrone on mental health. We present a case of dependence and psychosis in a patient using mephedrone (4-methylmethcathinone). The patient needed inpatient hospital care, was treated with antipsychotic olanzapine and recovered well.

Cyclic loading of rotator cuff repairs: A comparison of bioabsorbable tacks with metal suture anchors and transosseous sutures.

PURPOSE: The purposes of the study were (1) to compare rotator cuff repair strengths after cyclic loading of 2 bioabsorbable nonsuture-based tack-type anchors, transosseous sutures, and a metal suture-based anchor, and (2) to correlate bone mineral density with mode of failure and cycles to failure. We hypothesized that specimens with a lower bone density would fail through bone at a lower number of cycles independent of the method of cuff fixation. TYPE OF STUDY: Ex vivo biomechanical study. METHODS: Standardized full-thickness rotator cuff defects were created in 30 fresh-frozen cadaveric shoulders that were randomized to 1 of 4 repair groups: transosseous sutures; Mitek Super suture anchors (Mitek Surgical Products, Westwood, MA); smooth bioabsorbable 8-mm Suretacs (Acufex, Smith & Nephew Endoscopy, Mansfield, MA); or spiked bioabsorbable 8-mm Suretacs (Acufex). All repairs were cyclically loaded from 10 to 180 N; the numbers of cycles to 50% (gap, 5 mm) and 100% (gap, 10 mm) failure were recorded. RESULTS: In comparing the repair groups, we found only 1 significant difference: the number of cycles to 100% failure was significantly higher (P <.05) for the smooth bioabsorbable tack than for the transosseous suture group. There were no statistically significant (P

The use of carbon marking after stereotactic large-core-needle breast biopsy.

PURPOSE: To investigate the use of activated charcoal to mark the biopsy site and needle track after large-core-needle breast biopsy. MATERIALS AND METHODS: Three hundred seventy-six consecutive patients (with 383 lesions) were referred for stereotactic breast biopsy. Two hundred forty-seven lesions were carbon marked when the need for surgery was likely. Patients who underwent marking were followed up for the results of surgery or mammography performed at our institution. Specimen sizes obtained by using the carbon mark were compared with sizes of consecutive biopsy specimens obtained after hook-wire localization. RESULTS: Carbon marking was well tolerated in all cases. All 132 surgeries performed at the authors' institution were successful in removing the marked target. Specimen sizes compared favorably with sizes of comparison hook-wire localization specimens. All 68 lesions followed mammographically revealed no changes that were attributable to the use of carbon. Two minor complications were observed. Two small cancers were completely removed at needle biopsy. CONCLUSION: Carbon marking is safe and effective for marking the biopsy site and needle track created by stereotactic large-core-needle biopsy of the breast. Marking eliminates the need for postprocedural needle localization. It remains effective when small lesions have been completely removed. This technique should be considered in properly selected cases by those performing large-core-needle biopsy of the breast.

Popliteal pseudoaneurysm and arteriovenous fistula after arthroscopic meniscectomy.

A 69-year-old woman developed 2 pseudoaneurysms and an arteriovenous fistula in the popliteal vasculature after arthroscopic knee surgery. Despite heightened suspicion, prompt diagnosis, and early surgical intervention, the patient had an incomplete resolution of her symptoms.

Conformations and pharmacophores of cyclic RGD containing peptides which selectively bind integrin alpha(v)beta3.

This paper reports a detailed conformational characterization in solution by 1H-NMR in H2O and DMSO-d6 and molecular modeling simulations of cyclic peptides containing the RGDDV pharmacophore and the RGDY(Me)R pharmacophore. These two pentapeptide sequences when properly constrained in cyclic peptides are low to sub-nanomolar inhibitors of integrin alpha(v)beta3. The peptides containing the RGDDY(Me)R sequence bind potently to integrin alphaIIb3 as well. The conformations found in H2O and in DMSO-d6 solutions are valuable for the design of peptidomimetics of these two pharmacophores. The structure-activity relationships of the RGDDV and RGDY(Me)R pharmacophores within cyclic peptides are discussed. Specifically, the orientation of surface-accessible chemical features on the ligand, such as hydrophobic, positive and negative ionizable groups, which are considered to be responsible for the desired biological activity, is focused on.

Gene disruption studies of penicillin-binding proteins 1a, 1b, and 2a in Streptococcus pneumoniae.

The effects of inactivation of the genes encoding penicillin-binding protein 1a (PBP1a), PBP1b, and PBP2a in Streptococcus pneumoniae were examined. Insertional mutants did not exhibit detectable changes in growth rate or morphology, although a pbp1a pbp1b double-disruption mutant grew more slowly than its parent did. Attempts to generate a pbp1a pbp2a double-disruption mutant failed. The pbp2a mutants, but not the other mutants, were more sensitive to moenomycin, a transglycosylase inhibitor. These observations suggest that individually the pbp1a, pbp1b, and pbp2a genes are dispensable but that either pbp1a or pbp2a is required for growth in vitro. These results also suggest that PBP2a is a functional transglycosylase in S. pneumoniae.

Surface roughness enhances upward migration of bacteria on polymer fibers above liquid cultures.

Monofilament polypropylene (PP) fibers, very similar to fibers that have been used as monofilament tailstrings of interuterine contraceptive devices, were suspended vertically in bacterial liquid monocultures so that a portion of a fiber extended above the liquid surface. In some cases these highly oriented, cold drawn fibers were abraded prior to insertion in the cultures in order to produce surface roughness characterized by axial channels and protruding microfibrils that partially peeled from the fiber surface thereby forming the channels. Extent of migration on a fiber was assessed by aseptically cutting it into small segments, followed by culturing each segment on agar containing growth medium. Such assessment of the PP fibers after 48 h of incubation in the cultures revealed upward migration of Eschericia coli, Pseudomonas aeruginosa, and Staphylococcus aureus over significantly longer distances on the pre-roughened fibers than on those not so pre-treated. Mean measured distances of migration during 48 h were: for E. coli 2.7+/-0.6 mm on roughened fibers (n = 16) and 0.4+/-0.7 mm on fibers not roughened (n = 17); for S. aureus 9.0+/-4.3 mm on roughened fibers (n = 13) and 0.2+/-0.3 mm on fibers not roughened (n = 14); for P. aeruginosa 8.5+/-3.7 mm on roughened fibers (n = 26) and 0.2+/-0.5 mm on fibers not roughened (n = 5). Although no statistically significant (95% confidence level) difference could be discerned between the migration distances of S. aureus and P. aeruginosa, each of these species migrated a greater distance on the PP than did E. coli. The migrations observed are attributed predominantly to wicking of the liquid cultures upward in the axial grooves developed on the surface of the PP by the eruption and peeling of microfibrils from the surface. Surface tension of the growth medium was significantly lower than that of water and its contact angle on PP was less than 90 deg, thereby indicating a tendency to wet the PP. Bacterial growth in the medium further reduced its contact angle on PP, thereby indicating an even greater tendency to wet PP after such growth.

Novel glycopeptide antibiotics: N-alkylated derivatives active against vancomycin-resistant enterococci.

LY264826 (A82846B) is a naturally-occurring glycopeptide antibiotic, differing from vancomycin in the stereochemistry of the amino-sugar of the disaccharide function, and the presence of a third sugar attached at the benzylic position of amino acid residue 6. Despite these seemingly subtle differences, LY264826 is approximately 10 times more active than vancomycin against the enterococci. In the pursuit of new antibiotics active against multiresistant Gram-positive organisms, an extensive side chain SAR was developed focusing on the reductive alkylation of LY264826 at the amino function of the disaccharide moiety. A new series of derivatives having varying degrees of structural diversity in the side chain (e.g. varying lengths and degrees of rigidity) was found to have potent activity against vancomycin-resistant enterococci (MIC's < 1.0 microgram/ml) as well as activity against staphylococci and streptococci as good or better than vancomycin.

Enzymatic deacylation of teicoplanin followed by reductive alkylation: synthesis and antibacterial activity of new glycopeptides.

Novel glycopeptides derived from teicoplanin were prepared and evaluated for activity against antibiotic-resistant gram-positive pathogens. Removal of the fatty acid sidechains of teicoplanin was accomplished by enzymatic deacylation. The resulting deacylated teicoplanin was subjected to reductive alkylation resulting in mono- and di-alkylated compounds at the 2 possible primary amines. Deacylated teicoplanin was less active than teicoplanin against enterococci and staphylococci (MIC > or =32 microg/ml). All mono- and di-alkylated products regained some activity, and some had potent activity against both staphylococci and glycopeptide-resistant enterococci. MICs of the most potent di-alkylated compounds ranged from 0.25 approximately 2 microg/ml against glycopeptide-resistant enterococci.

Semisynthetic glycopeptide antibiotics derived from LY264826 active against vancomycin-resistant enterococci.

Certain derivatives of the glycopeptide antibiotic LY264826 with N-alkyl-linked substitutions on the epivancosamine sugar are active against glycopeptide-resistant enterococci. Six compounds representing our most active series were evaluated for activity against antibiotic-resistant, gram-positive pathogens. For Enterococcus faecium and E. faecalis resistant to both vancomycin and teicoplanin, the MICs of the six semisynthetic compounds for 90% of the strains tested were 1 to 4 micrograms/ml, compared with 2,048 micrograms/ml for vancomycin and 256 micrograms/ml for LY264826. For E. faecium and E. faecalis resistant to vancomycin but not teicoplanin, the MICs were 0.016 to 1 micrograms/ml, compared with 64 to 1,024 micrograms/ml for vancomycin. The compounds were highly active against vancomycin-susceptible enterococci and against E. gallinarum and E. casseliflavus and showed some activity against isolates of highly vancomycin-resistant leuconostocs and pediococci. The MICs for 90% of the strains of methicillin-resistant Staphylococcus aureus tested were typically 0.25 to 1 micrograms/ml, compared with 1 microgram/ml for vancomycin. Against methicillin-resistant S. epidermidis MICs ranged from 0.25 to 2 micrograms/ml, compared with 1 to 4 micrograms/ml for vancomycin and 4 to 16 micrograms/ml for teicoplanin. The spectrum of these new compounds included activity against teicoplanin-resistant, coagulase-negative staphylococci. The compounds exhibited exceptional potency against pathogenic streptococci, with MICs of < or = 0.008 microgram/ml against Streptococcus pneumoniae, including penicillin-resistant isolates. In in vivo studies with a mouse infection model, the median effective doses against a challenge by S. aureus, S. pneumoniae, or S. pyogenes were typically 4 to 20 times lower than those of vancomycin. Overall, these new glycopeptides, such as LY307599 and LY333328, show promise for use as agents against resistant enterococci, methicillin-resistant S. aureus, and penicillin-resistant pneumococci.

Reductive alkylation of glycopeptide antibiotics: synthesis and antibacterial activity.

Reductive alkylation of the A82846 family of glycopeptide antibiotics has the potential of producing seven products. N-Alkylation of the disaccharide amino function can be accomplished selectively, and offers the greatest increase in antibacterial activity. Products resulting from N-alkylation of LY264826 (A82846B) provide the most potent derivatives as compared to other members of this class of antibiotics. Two of these derivatives, LY307599 and LY333328 are approximately 500 times more active than vancomycin against vancomycin-resistant enterococci.