Michael W. Hughes

The mutagenic potential of duodenoesophageal reflux.

Summary Background Data:Duodenogastric–esophageal reflux disease is directly linked to Barretts esophagus and to the development of esophageal adenocarcinoma. Despite this link, little is known about the mutagenic potential of refluxed material on the esophageal mucosa. We hypothesize that the reflux of gastric and duodenal content causes mutations in esophageal mucosa in vivo. Methods:Seven Sprague Dawley/Big Blue F1 lacI transgenic rats underwent esophagoduodenostomy (ED) to surgically create duodeno–gastric–esophageal reflux. Fourteen nonoperated rats served as negative (n = 7) and as positive (n = 7/methyl-N-amyl-nitrosamine [MNAN] intraperitoneally) controls. The animals were killed 16 weeks after operation or injection, the entire esophageal mucosa was harvested, and mutation frequency was determined through standard Big Blue Mutagenesis Assay. Results:Gross esophagitis was evident in all operated animals. The frequency of lacI mutations in esophageal mucosal cells of animals with ED was significantly higher, nearly 1.5-fold, than that of nonoperated animals. Nitrosamine administration resulted in a nearly 20-fold increase of lacI mutation frequency. Thirteen mutations were successfully sequenced, 46% occurred at CpG dinucleotide sites and 61% were either C to T or G to A transitions. Conclusions:The data provide preliminary evidence of the mutagenic potential of bile reflux on esophageal epithelium. The specific mutations are markedly higher than would be expected by chance and are similar to that found in p53 mutations of human esophageal adenocarcinoma, providing a link to human esophageal cancer.

Systematic review and meta-analysis of continuous local anaesthetic wound infiltration versus epidural analgesia for postoperative pain following abdominal surgery

Local anaesthetic wound infiltration techniques reduce opiate requirements and pain scores. Wound catheters have been introduced to increase the duration of action of local anaesthetic by continuous infusion. The aim was to compare these infiltration techniques with the current standard of epidural analgesia.

Endoscopic augmentation of the cardia with a biocompatible injectable polymer (Enteryx) in a porcine model

Background: Endoscopic approaches to restore the gastroesophageal barrier in patients with gastroesophageal reflux disease (GERD) are presently undergoing clinical trial. The aim of the study was to demonstrate the feasibility, durability, safety, and antireflux efficacy following augmentation of the cardia with a biocompatible injectable polymer (Enteryx). Methods: Augmentation was performed in 12 Yucatan mini-pigs. The cardia was injected circumferentially with 1–1.5 ml of Enteryx at three or four sites. Four groups of three animals each were killed at 2, 6, 12, and 24 weeks following augmentation. Gastrointestinal endoscopy and esophageal manometry were performed preoperatively and postoperatively. Competency was determined as the intragastric pressure (yield pressure) and volume (yield volume) needed during gastric distension with air and water to result in equalization of gastric and esophageal pressure. Comparisons were made with a group of noninjected animals (n = 6). Results: All animals had a normal eating pattern; none showed any evidence of vomiting or regurgitation. The median injection volume was 4 ml (range, 1–8). At autopsy, implants were found in 83% of the animals. Intramuscular placement of the implant was durable, whereas sloughing occurred if the implant was placed submucosally. The mechanical properties of sphincter length and pressure were unaffected by the injection. The median yield pressure of the animals that survived for >6 weeks (21.4 mmHg) was significantly greater (p = 0.049) than the animals that survived for <6 weeks (4.5 mmHg) and greater (p = 0.054) than the control animals (9.1 mmHg), suggesting that the healing process was associated with reduced distensibility of the cardia. Conclusions: Augmentation of the cardia with an injectable polymer (Enteryx) is simple, safe, and durable. Early studies suggest that alteration in the distensibility and geometry of the gastroesophageal junction may provide antireflux protection.

Organ-Level Quorum Sensing Directs Regeneration in Hair Stem Cell Populations

Coordinated organ behavior is crucial for an effective response to environmental stimuli. By studying regeneration of hair follicles in response to patterned hair plucking, we demonstrate that organ-level quorum sensing allows coordinated responses to skin injury. Plucking hair at different densities leads to a regeneration of up to five times more neighboring, unplucked resting hairs, indicating activation of a collective decision-making process. Through data modeling, the range of the quorum signal was estimated to be on the order of 1 mm, greater than expected for a diffusible molecular cue. Molecular and genetic analysis uncovered a two-step mechanism, where release of CCL2 from injured hairs leads to recruitment of TNF-α-secreting macrophages, which accumulate and signal to both plucked and unplucked follicles. By coupling immune response with regeneration, this mechanism allows skin to respond predictively to distress, disregarding mild injury, while meeting stronger injury with full-scale cooperative activation of stem cells.

Progressive Alopecia Reveals Decreasing Stem Cell Activation Probability during Aging of Mice with Epidermal Deletion of DNA Methyltransferase 1

To examine the roles of epigenetic modulation on hair follicle regeneration, we generated mice with a K14Cre-mediated loss of DNA methyltransferase (DNMT) 1. The mutant shows an uneven epidermal thickness and alterations in hair follicle size. When formed, hair follicle architecture and differentiation appear normal. Hair subtypes exist but hair fibers are shorter and thinner. Hair numbers appear normal at birth but gradually decrease to fewer than 50% of control in 1 year old mice. Sections of old mutant skin shows follicles in prolonged telogen with hyperplastic sebaceous glands. Anagen follicles in mutants exhibit decreased proliferation and increased apoptosis in matrix transient amplifying cells. Although K15 positive stem cells in the mutant bulge are comparable in number to the control, their ability to proliferate and become activated to form a hair germ is reduced. As mice age, residual DNMT activity declines further and the probability of successful anagen reentry decreases, leading to progressive alopecia. Paradoxically, there is increased proliferation in the epidermis which also shows aberrant differentiation. These results highlight the importance of DNA methylation in maintaining stem cell homeostasis during the development and regeneration of ectodermal organs.

Therapeutic strategy for hair regeneration: hair cycle activation, niche environment modulation, wound-induced follicle neogenesis, and stem cell engineering.

Introduction: There are major new advancements in the fields of stem cell biology, developmental biology, regenerative hair cycling, and tissue engineering. The time is ripe to integrate, translate, and apply these findings to tissue engineering and regenerative medicine. Readers will learn about new progress in cellular and molecular aspects of hair follicle development, regeneration, and potential therapeutic opportunities these advances may offer. Areas covered: Here, we use hair follicle formation to illustrate this progress and to identify targets for potential strategies in therapeutics. Hair regeneration is discussed in four different categories: i) Intra-follicle regeneration (or renewal) is the basic production of hair fibers from hair stem cells and dermal papillae in existing follicles. ii) Chimeric follicles via epithelial–mesenchymal recombination to identify stem cells and signaling centers. iii) Extra-follicular factors including local dermal and systemic factors can modulate the regenerative behavior of hair follicles, and may be relatively easy therapeutic targets. iv) Follicular neogenesis means the de novo formation of new follicles. In addition, scientists are working to engineer hair follicles, which require hair-forming competent epidermal cells and hair-inducing dermal cells. Expert opinion: Ideally self-organizing processes similar to those occurring during embryonic development should be elicited with some help from biomaterials.

Attitudes of patients and care providers to enhanced recovery after surgery programs after major abdominal surgery

BACKGROUND Enhanced recovery after surgery (ERAS) is a well-established pathway of perioperative care in surgery in an increasing number of specialties. To implement protocols and maintain high levels of compliance, continued support from care providers and patients is vital. This survey aimed to assess the perceptions of care providers and patients of the relevance and importance of the ERAS targets and strategies. MATERIALS AND METHODS Pre- and post-operative surveys were completed by patients who underwent major hepatic, colorectal, or oesophagogastric surgery in three major centers in Scotland, Norway, and The Netherlands. Anonymous web-based and article surveys were also sent to surgeons, anesthetists, and nurses experienced in delivering enhanced recovery protocols. Each questionnaire asked the responder to rate a selection of enhanced recovery targets and strategies in terms of perceived importance. RESULTS One hundred nine patients and 57 care providers completed the preoperative survey. Overall, both patients and care providers rated the majority of items as important and supported ERAS principles. Freedom from nausea (median, 10; interquartile range [IQR], 8-10) and pain at rest (median, 10; IQR, 8-10) were the care components rated the highest by both patients and care providers. Early return of bowel function (median, 7; IQR, 5-8) and avoiding preanesthetic sedation (median, 6; IQR, 3.75-8) were scored the lowest by care providers. CONCLUSIONS ERAS principles are supported by both patients and care providers. This is important when attempting to implement and maintain an ERAS program. Controversies still remain regarding the relative importance of individual ERAS components.

Accelerated closure of skin wounds in mice deficient in the homeobox gene Msx2.

Differences in cellular competence offer an explanation for the differences in the healing capacity of tissues of various ages and conditions. The homeobox family of genes plays key roles in governing cellular competence. Of these, we hypothesize that Msx2 is a strong candidate regulator of competence in skin wound healing because it is expressed in the skin during fetal development in the stage of scarless healing, affects postnatal digit regeneration, and is reexpressed transiently during postnatal skin wound repair. To address whether Msx2 affects cellular competence in injury repair, 3 mm full‐thickness excisional wounds were created on the back of C.Cg‐Msx2tm1Rilm/Mmcd (Msx2 null) mice and the healing pattern was compared with that of the wild type mice. The results show that Msx2 null mice exhibited faster wound closure with accelerated reepithelialization plus earlier appearance of keratin markers for differentiation and an increased level of smooth muscle actin and tenascin in the granulation tissue. In vitro, keratinocytes of Msx2 null mice exhibit increased cell migration and the fibroblasts show stronger collagen gel contraction. Thus, our results suggest that Msx2 regulates the cellular competence of keratinocytes and fibroblasts in skin injury repair.

In search of the Golden Fleece: Unraveling principles of morphogenesis by studying the integrative biology of skin appendages

The mythological story of the Golden Fleece symbolizes the magical regenerative power of skin appendages. Similar to the adventurous pursuit of the Golden Fleece by the multi-talented Argonauts, today we also need an integrated multi-disciplined approach to understand the cellular and molecular processes during development, regeneration and evolution of skin appendages. To this end, we have explored several aspects of skin appendage biology that contribute to the Turing activator/inhibitor model in feather pattern formation, the topo-biological arrangement of stem cells in organ shape determination, the macro-environmental regulation of stem cells in regenerative hair waves, and potential novel molecular pathways in the morphological evolution of feathers. Here we show our current integrative biology efforts to unravel the complex cellular behavior in patterning stem cells and the control of regional specificity in skin appendages. We use feather/scale tissue recombination to demonstrate the timing control of competence and inducibility. Feathers from different body regions are used to study skin regional specificity. Bioinformatic analyses of transcriptome microarrays show the potential involvement of candidate molecular pathways. We further show Hox genes exhibit some region specific expression patterns. To visualize real time events, we applied time-lapse movies, confocal microscopy and multiphoton microscopy to analyze the morphogenesis of cultured embryonic chicken skin explants. These modern imaging technologies reveal unexpectedly complex cellular flow and organization of extracellular matrix molecules in three dimensions. While these approaches are in preliminary stages, this perspective highlights the challenges we face and new integrative tools we will use. Future work will follow these leads to develop a systems biology view and understanding in the morphogenetic principles that govern the development and regeneration of ectodermal organs.

Non-invasive evaluation of therapeutic response in keloid scar using diffuse reflectance spectroscopy.

The pathogenesis and ideal treatment of keloid are still largely unknown, and it is essential to develop an objective assessment of keloid severity to evaluate the therapeutic response. We previously reported that our diffuse reflectance spectroscopy (DRS) system could assist clinicians in understanding the functional and structural condition of keloid scars. The purpose of this study was to understand clinical applicability of our DRS system on evaluating the scar severity and therapeutic response of keloid. We analyzed 228 spectral data from 71 subjects with keloid scars. The scars were classified into mild (0-3), moderate (4-7) and severe (8-11) according to the Vancouver scar scale. We found that as the severity of the scar increased, collagen concentration and water content increased, and the reduced scattering coefficient at 800 nm and oxygen saturation (SaO2) decreased. Using the DRS system, we found that collagen bundles aligned in a specific direction in keloid scars, but not in normal scars. Water content and SaO2 may be utilized as reliable parameters for evaluating the therapeutic response of keloid. In conclusion, the results obtained here suggest that the DRS has potential as an objective technique with which to evaluate keloid scar severity. In addition, it may be useful as a tool with which to track longitudinal response of scars in response to various therapeutic interventions.