The impact of home enteral feeding on the daily lives of people with head and neck cancer: a metasynthesis of qualitative studies.

BACKGROUND: Home enteral feeding (HEF) may have a wider impact on peoples' daily lives beyond influencing their nutritional and clinical status. This metasynthesis aimed to determine the impact of HEF on peoples' daily lives. METHODS: Qualitative studies were included with adults, who had been diagnosed with head and neck cancer and had finished their cancer treatment, with a feeding tube in place. Medline, PubMed and Cinahl were searched (August 2009 to August 2019). Thematic synthesis was conducted to interpret findings from the included studies. The 'Confidence in the Evidence from Reviews of Qualitative research' (GRADE-CERQual) approach was used to assess the level of confidence associated with each review finding. RESULTS: Seven qualitative studies met the eligibility criteria. Four overarching themes were identified. 'Loss of life as they once knew it' encompassed loss of normality. 'Developing personal coping strategies works towards restoring a sense of normality' encapsulated active adjustment and acceptance of the feeding tube. 'Navigating the hurdles when transitioning back to eating' comprised the trials and tribulations of returning to oral intake. Overall, participants recognised that they could not have managed without the feeding tube and this is encapsulated in 'Feeding tube valued'. CONCLUSIONS: In many cases, initial feelings of change and loss as a result of HEF were replaced with empowerment, adaptation and acceptance following a period of adjustment. Despite the challenges associated with HEF, participants acknowledged its purpose as being functional, as well as reassuring. Future research should explore barriers and facilitators to self-management and patient empowerment amongst those receiving HEF.

Short bowel syndrome and the impact on patients and their families: a qualitative study.

BACKGROUND: Short bowel syndrome (SBS) is a malabsorptive condition that can result in intestinal failure (SBS-IF). Many patients with SBS-IF require home parenteral nutrition (PN) for survival. However, PN has profound effects on patients and their family members. The present study aimed to understand the lived experience of SBS-IF for patients and their families. METHODS: In-depth semi-structured qualitative interviews were conducted with 15 patients with SBS-IF and five adult family members living with someone with SBS-IF. A patient-centric approach was taken, with a patient steering group providing input and guidance to develop the interview guide. Key concepts were identified using thematic analysis of interview transcripts. RESULTS: Patients' lives were dominated by having SBS-IF. They described physical impacts that included patient-reported signs and symptoms and physical restrictions comprising of restrictions on daily life, actives of daily living and physical functioning. In addition, they encountered emotional impacts with a plethora of negative feelings and social impacts, such as difficulties socialising and maintaining relationships. Patients coped by adapting their life around SBS-IF, having support and adopting an attitude of gratitude and acceptance. Family members were also affected and, along with patients, appreciated the respite of a night off from infusions. CONCLUSIONS: Patients and families face many difficulties with SBS-IF. Healthcare professionals can support patients by facilitating them explore what others have found beneficial; adapting their life around PN, viewing PN with acceptance and trying to cultivate gratitude. Further research into the support required for families may be beneficial.

Dealing with loss: food and eating in women with ovarian cancer on parenteral nutrition.

BACKGROUND: Malignant bowel obstruction is a common complication of ovarian cancer, resulting in limited oral intake. Home parenteral nutrition (HPN) may be offered to patients in this condition to meet nutritional requirements. However, it is not known how they experience being unable to eat. The present study reports how patients related to food when receiving HPN. METHODS: The investigation was a qualitative study underpinned by phenomenology with women with advanced ovarian cancer in bowel obstruction receiving parenteral nutrition. Interview transcripts were analysed thematically guided by the techniques of Van Manen. RESULTS: We recruited 20 women to the study. Participants were interviewed a maximum of four times and a total of 39 in-depth longitudinal interviews were conducted. Participants could tolerate minimal amounts of food, if they had a venting gastrostomy. Not being able to eat engendered a sense of sadness and loss, and most women found it challenging to be in the presence of others eating. They adopted strategies to cope, which included fantasising about food and watching cookery programmes. These approaches were not a long-term solution; either participants came to terms with their loss or the strategies became less effective in providing relief. CONCLUSIONS: Home parenteral nutrition meets the nutritional requirements of patients with malignant bowel obstruction but cannot replace the non-nutritive functions of food. Healthcare professionals can offer a patient-centred approach by acknowledging the difficulties that patients may face and, wherever possible, encourage them to focus on the positive benefits of interacting with people rather than the loss of eating on social occasions.

Priority setting for adult malnutrition and nutritional screening in healthcare: a James Lind Alliance.

BACKGROUND: Malnutrition is one the greatest global health challenges of our generation, leading to the increased utilisation of healthcare resources, as well as morbidity and mortality. Research has primarily been driven by industry, academia and clinical working groups and has had little involvement from patients and carers. The project described in the present study aimed to establish a priority setting partnership allowing patients, carers and healthcare professionals an opportunity to influence the research agenda. METHODS: A national survey was conducted to gather malnutrition uncertainties and identify key issues (i.e. areas within scope where an evidence-base is lacking) from those with experience of malnutrition. Uncertainties were analysed according to themes. Similar questions were grouped and summary questions were developed. A second survey was conducted and respondents were asked to choose their 10 most important summary questions. A workshop was conducted to finalise the top 10 research priorities from the most frequently indicated uncertainties on the interim survey. RESULTS: Overall, 1128 uncertainty questions were submitted from 268 people. The interim survey had 71 responses and a list of the top 26 questions was generated for the workshop. There were 26 questions discussed, ranked and agreed by healthcare professionals, carers and patients at the workshop. The top 10 research priorities were then chosen. These included questions on oral nutritional supplements, vulnerable groups, screening, community care, use of body mass index and technology. CONCLUSIONS: The top 10 research priorities in malnutrition and nutritional screening have been identified from a robust process involving patients, carers and healthcare professionals.

The impact of living with home enteral feeding: perspectives of people who have had a diagnosis of head and neck cancer.

BACKGROUND: The number of people with head and neck cancer who are home enterally fed continues to grow each year. Insertion of a feeding tube is common place in these patients and is considered to have a detrimental effect on quality of life. The present study aimed to investigate the daily impact of home enteral feeding (HEF) from the perspective of people who have had a diagnosis of head and neck cancer. METHODS: The methodology aligned with interpretative phenomenology analysis. People who were home enterally fed, with head and neck cancer, and aged ≥18 years were recruited. Data were collected using semi-structured interviews and analysis focused on what the daily impact of HEF meant for participants. RESULTS: Data saturation was achieved after interviewing 15 participants. Five cluster themes were identified. 'Deviation from the norm' encompassed change and loss of normality. 'Regaining control leading to empowerment' encompassed participant empowerment through development of new skills and adjusting the feeding regime. 'Creating a new normal' involved making adjustments to facilitate inclusion and participation. 'External modifiers of the HEF experience' and 'internal modifiers of the HEF experience' encompassed the identification of external and internal HEF factors that influenced HEF adaptation. CONCLUSIONS: HEF was found to influence peoples' daily lives substantially and required extensive adjustments for individuals to find a new normal. A greater level of interpretation was provided beyond the current evidence-base for this group. Policymakers and clinicians should recognise the wider impact of HEF and ensure that this awareness is embedded in clinical practice.

Healthy eating interventions in adults living with and beyond colorectal cancer: a systematic review.

BACKGROUND: Rates of cancer survival are increasing, with more people living with and beyond cancer. Lifestyle recommendations for cancer survivors are based largely on extrapolation from cancer prevention recommendations. The present study aimed to systematically review the literature on randomised controlled trials (RCTs) focusing on healthy eating interventions in people with colorectal cancer (CRC). METHODS: A structured search of electronic databases was conducted in March 2018 using medical subject headings (MeSH) and text words related to CRC and diet. The results of the literature searches were uploaded to online software for data management. Titles and abstracts were screened based on the inclusion and exclusion criteria and data were extracted. Quality of data was assessed using the Cochrane Handbook. RESULTS: Seven studies were identified, including six RCTs and one RCT protocol, with a total of 2233 participants from six studies, of whom 1010 (45%) had CRC. Three studies assessed anthropometrics demonstrating participants who received dietary intervention had a greater reduction in measurements. Six studies assessed changes in dietary components; however, only one demonstrated an increase in dietary fibre. Two studies reported improvements in quality of life favouring dietary intervention groups. CONCLUSIONS: The quality of identified studies was variable, with limited evidence to support dietary intervention improving dietary intake in people living with or after CRC. Studies to date have not been based on robust study design that has combined all dietary interventions linked to CRC. As a result of the heterogeneity of the studies identified, it was difficult to draw strong conclusions.

Practical measurement of body composition using bioelectrical impedance, air displacement plethysmography and ultrasound in stable outpatients with short bowel syndrome receiving home parenteral nutrition: comparison of agreement between the methods.

BACKGROUND: People diagnosed with intestinal failure (IF) as a result of short bowel syndrome are dependent on home parenteral nutrition (HPN). Measuring nutritional status is essential for monitoring treatment. The present study aimed to determine the agreement and feasibility of three methods bioelectrical impedance analysis (BIA), ultrasound and air displacement plethysmography (ADP) for measuring body composition in people receiving HPN. METHODS: Body composition data were collected from patients attending an IF clinic. RESULTS: There were 50 participants recruited and data were collected for BIA (n = 46), ultrasound (n = 49) and ADP (n = 9). Numbers for ADP were much lower because of a lack of participant uptake. Fat-free mass (FFM) measured by BIA and ultrasound in comparison to ADP was found to have good intraclass correlation (ICC) 0.791 (95% confidence interval (CI) CI  -0.21 to 0.96) and a moderate ICC 0.659 [95% (CI) -0.27 to 0.92], respectively. Fat mass (FM) measured by both BIA and ultrasound in comparison to ADP was found to have moderate ICC 0.660 (95% CI -0.28 to 0.92) and poor ICC -0.005 (95% CI -0.73 to 0.65), respectively. CONCLUSIONS: Compared to ADP, BIA indicated moderate to good agreement for measuring body composition, whereas ultrasound indicated far less agreement, particularly when measuring FM. The lack of uptake of ADP suggests that participants found the Bodpod (COSMED Srl, Shepperton, UK) unfavourable. Considering that ultrasound has limited agreement and ADP was not the preferred option for participants, BIA shows some potential. However, the difference between ADP and BIA was larger for FM compared to FFM, which needs to be considered in the clinical setting.

A systematic review of the use of ketogenic diets in adult patients with cancer.

BACKGROUND: A growing body of evidence indicates the importance of nutrition in cancer treatment. Ketogenic diets are one strategy that has been proposed to enhance traditional anticancer therapy. This review summarises the evidence concerning the effect of oral ketogenic diets on anthropometry, metabolism, quality of life (QoL) and tumour effects, at the same time as documenting adverse events and adherence in patients with cancer. METHODS: We searched electronic databases using medical subject headings (MeSH) and text words related to ketogenic diets and cancer. Adult patients following a ketogenic diet as a complementary therapy prior, alongside or after standard anticancer treatment for more than 7 days were included. Studies were assessed for quality using the Critical Appraisal Skills Programme tools (https://www.casp-uk.net). RESULTS: Eleven studies were included with 102 participants (age range 34-87 years) from early-phase trials, cohort studies and case reports. Studies included participants with brain, rectal or mixed cancer sites at an early or advanced disease stage. The duration of intervention ranged from 2.4 to 134.7 weeks (0.5-31 months). Evidence was inconclusive for nutritional status and adverse events. Mixed results were observed for blood parameters, tumour effects and QoL. Adherence to diet was low (50 out of 102; 49%) and ranged from 23.5% to 100%. CONCLUSIONS: High-quality evidence on the effect of ketogenic diets on anthropometry, metabolism, QoL and tumour effects is currently lacking in oncology patients. Heterogeneity between studies and low adherence to diet affects the current evidence. There is an obvious gap in the evidence, highlighting the need for controlled trials to fully evaluate the intervention.

Patient experiences of perioperative nutrition within an Enhanced Recovery After Surgery programme for colorectal surgery: a qualitative study.

AIM: Nutrition is an important element of the Enhanced Recovery After Surgery (ERAS) programme. Patients have previously indicated that nutrition is a key component of ERAS that requires improvement. Our aim was to explore the perioperative nutrition experiences of colorectal surgical patients to identify barriers and facilitators to the integration of nutrition within ERAS. METHOD: Sixteen individuals undergoing colorectal surgery participated in a semi-structured interview between postoperative day three and hospital discharge. The topic guide was developed iteratively throughout the study; topics included preoperative counselling, carbohydrate loading, fasting and postoperative nutrition. A constant comparison technique was employed during coding, and an inductive thematic analysis was used. Validity was ensured by double coding a sample of transcripts. RESULTS: Findings are presented in the context of the following clinical themes: preoperative information, preoperative fasting, carbohydrate loading and nutritional drinks, postoperative diet and discharge. Individuals received too much general information which was repetitive, contradictory and not disease specific; this formed a key barrier affecting nutrition. Other barriers were negative experiences of nutritional drinks, stoma management, nausea and vomiting, and challenges from the hospital environment. Facilitators included interactions with staff, food accessibility and choice, and motivation for discharge. CONCLUSION: The key barrier to adherence of perioperative nutrition protocols was poor provision of information. Targeted information regarding postoperative diet, stoma management and coping with nausea and vomiting would be beneficial for colorectal surgical patients. Easily accessible food provided by ward staff was considered a facilitator.

An exploration of food and the lived experience of individuals after treatment for colorectal cancer using a phenomenological approach.

BACKGROUND: There is a paucity of qualitative literature investigating people's experiences of food and nutrition after treatment for cancer. The present study aimed to explore people's relationships with food and nutrition throughout their colorectal cancer journey. METHODS: In-depth semi-structured interviews were conducted with 25 participants who had undergone surgery for colorectal cancer. The study design was informed by principles of phenomenology. Data were collected then transcribed and analysed using an inductive coding process and a thematic analysis to allow the themes to highlight people's lived experiences. RESULTS: Data enabled five primary themes to be drawn including: 'appetite swings', 'emotions on a changing physicality', 'the medicalisation of food', 'taking control of symptom management' and a cross-cutting theme 'drivers and vehicles for action'. Feelings and emotions described by participants around their relationship with food and nutritional status often guided decisions on what was eaten more than objective nutritional measure or dietary advice. Participants used weight changes, appetite and food as barometers to measure their overall recovery. Food was an area over which people exhibited control of their lives and they could quantify, in measurable units, their overall well-being and rehabilitation. They did this either by using the currency of body weight in pounds or the size of portions eaten. CONCLUSIONS: Appetite, weight and symptoms influenced dietary intake substantially and were poignant issues affecting people's lives. The relationship people have with food determines their eating habits and an understanding of the essences and nuances of their experiences is essential to enable the delivery of patient-centred care.

An explorative study of the views and experiences of food and weight loss in patients with operable pancreatic cancer perioperatively and following surgical intervention.

PURPOSE: Malnutrition and weight loss are highly prevalent in patients with pancreatic cancer. However, little is known about how patients experience such changes after completion of a surgery. The aim of this study was to explore the views and experiences about weight loss and food in patients with operable pancreatic cancer perioperatively and following surgical treatment. METHODS: An exploratory study underpinned by hermeneutic phenomenological philosophy was conducted. Thirteen post-surgery patients with pancreatic cancer were recruited from a National Health Service (NHS) Trust in the north of England. Sampling was purposive and data were collected using semi-structured in-depth interviews. Each interview was digitally recorded, transcribed verbatim and analysed using NVivo 8. RESULTS: Six themes emerged from the data that represented the patients' views and experience of foods and weight loss after surgery: "struggling with weight loss", "being pressured to eat", "experiences with nutrition support therapy", "perception of the role of the dietitians", "lacking appropriate dietary instructions" and "road to recovery". CONCLUSIONS: The study has found that sufficient dietary advice, appropriate nutrition support and patient self-management are significant factors that affect how people recover from surgery for pancreatic cancer. Further work is required to understand the nature and effect of weight loss in surgical patients with pancreatic cancer and the role of food in their weight management.

A systematic review and meta-analysis of the effects of community use of oral nutritional supplements on clinical outcomes.

The impact of oral nutritional supplements (ONS) on patients with complications (disease related morbidity) requires further exploration. This systematic review included 44 randomised controlled trials (RCT) (29 RCT surgical, 15 RCT medical patients) examining the effect of ONS in community settings on the incidence of complications (n = 716, mean age 67 years, range 35-87). ONS (mean intake 588 kcal/day, range 125-1750; protein 22 g/day, range 0-54; mean energy from protein 22 %, range 0-54) were prescribed for a mean 74 days, range 5-365. Most RCT (77 %) reported fewer complications in the ONS group versus control. Meta-analysis (39 RCT) showed ONS consumption reduced complications including infections, pressure ulcers, wound and fracture healing (OR 0.68, 95 % CI 0.59,0.79; p<0.001). Results showed reductions when ONS were used in hospital and community settings (OR 0.72, 95 % CI 0.59,0.87; p = 0.001) or just in the community (OR 0.65, 95 % CI 0.52, 0.80; p<0.001). Reductions in complications were only seen with high ONS adherence ≥ 80 % (OR 0.63, 95 % CI 0.48,0.83; p = 0.001) and ready-to-drink ONS (OR 0.69, 95 % CI 0.60,0.81; p<0.001). This systematic review and meta-analysis show community-based use of ONS in addition to the diet substantially reduces the incidence of complications. The diversity of ONS, patient populations and complication outcomes within the trials included in this review mean further research is warranted.

A multi-national survey of experience and attitudes towards commencing home parenteral nutrition for patients with advanced cancer.

INTRODUCTION: Advanced cancer (AC) is increasingly an indication for home parenteral nutrition (HPN) but an area with possible variation in practice between geographical locations. The aims of this study are to explore the views and experiences of international multi-disciplinary teams to determine opinions and practices. METHODS: An online questionnaire was developed with members of the Home Artificial Nutrition and Chronic Intestinal Failure interest group of the European Society for Clinical Nutrition and Metabolism (ESPEN) and distributed to colleagues involved in managing patients with AC on HPN. RESULTS: A total of 220 responses were included from 5 continents including 36 countries, with 90% of all responses from Europe. Predicted survival was a key factor influencing the decision to commence HPN for most respondents 152/220 (75%), with the majority of participants reporting that patients should have a predicted survival of ≥3 months if considered for HPN (≥3 months: n = 124, 56% vs. <3 months: n = 47, 21%, p < 0.001). However, most respondents were not confident about predicting overall survival in more than 50% of cases (confident n = 40, 23% vs not confident n = 135, 77%, p < 0.001). Barriers to utilising HPN in AC included colleagues' objections (n = 91, 46%), lack of local expertise (n = 55, 28%) and funding restrictions (n = 34, 17%). CONCLUSIONS: Significant consensus was observed regarding AC as indication for HPN, while areas of variation exist. Survival prognostication is often used as an indication for commencing HPN in people with AC, although the majority of respondents were not confident in prognosticating, suggesting better clinical prognostication tools will be of assistance. Further studies are also required to better understand the obstacles faced by clinical teams to commencing HPN that may explain variations in clinical practice between countries, as well as adressing variation in funding.

An unblinded randomised controlled trial of preoperative oral supplements in colorectal cancer patients.

BACKGROUND: Perioperative oral supplementation has been shown to reduce post-operative complications. However, the use of preoperative standard oral supplements in a cohort of colorectal cancer patients has not been evaluated. The present study examined whether preoperative supplements are beneficial in this group. METHODS: In a randomised controlled trial, patients were assigned to receive 400 mL of oral supplement and dietary advice or dietary advice alone. Primary outcome was the number of post-operative complications. One hundred and twenty-five patients were recruited (59 randomised to the intervention group and 66 to the control group) and nine were excluded. RESULTS: In the intervention group, 24 (44%) patients had a complication compared to 26 (42%) in the control group (P = 0.780). In the intervention and control groups, there were eight (15%) and 16 (25%) surgical site infections, respectively (P = 0.140) and seven (13%) and 11 (17%) chest infections, respectively (P = 0.470). Subgroup analysis for hypothesis generation included 83 (71%) weight-losing patients, where there was a significant reduction in surgical site infections using the Buzby definition (P = 0.034), although this was not the case for the Centre for Disease Control definition (P = 0.052). CONCLUSIONS: There was no evidence that preoperative supplements were beneficial in reducing the number of complications, although there may be some benefit for surgical site infections in selected weight-losing preoperative patients.

Nutritional status of preoperative colorectal cancer patients.

BACKGROUND: The present study aimed to determine the extent of malnutrition in preoperative colorectal cancer patients. Malnutrition has been shown to affect post-operative outcome, so it would be beneficial to identify those who are malnourished or who are at risk of becoming so preoperatively. We examine whether weight loss is related to the length of stay or changes in fat free mass. METHODS: Patients were enrolled consecutively from outpatients 2-4 weeks prior to surgery. Assessments included body mass index, percentage weight loss, dynamometry, Malnutrition Universal Screening Tool, Subjective Global Assessment and bioelectrical impedance. Cancer staging and hospital length of stay were recorded. RESULTS: One hundred and thirty-two patients were eligible and 87 enrolled. Sixty-seven patients were weight losing and 20% had lost >10% of their usual body weight. Handgrip strength was lower in malnourished patients compared to those who had not lost weight (mean 19.4 and 27.3 kg, respectively, P = 0.013). Mean (SD) fat free mass in patients with a weight loss >10% was 39.7 (13.5) kg and, in those with <10% weight loss, was 51.9 (12.0) kg (P = 0.001). This difference was not demonstrated for fat. CONCLUSIONS: Over half of these patients had lost weight prior to surgery and one in five were malnourished. Body composition measurements demonstrated that malnourished patients had significantly less fat free mass compared to patients who were not clinically malnourished. Nutritional screening would be beneficial in this group preoperatively to identify weight-losing patients at an early stage in the care pathway when they initially enter the secondary care system.

Identification of an intracellular postsynaptic antigen at the frog neuromuscular junction.

A layer of amorphous, electron-dense material is situated at the cytoplasmic surface of the postsynaptic membrane of vertebrate neuromuscular synapses. The function of this structure is not clear, but its location suggests that it may have an important role in the formation and/or maintenance of the synapse. This paper demonstrates that a monoclonal antibody raised against antigens from Torpedo electric organ binds to an intracellular, postsynaptic protein at the frog neuromuscular synapse. Indirect immunofluorescence on frozen sections of frog muscle was used to demonstrate that the antigen is concentrated at synaptic sites in normal muscle. In denervated muscle, the antigen remains concentrated at synaptic sites, but is also present at extrasynaptic regions of denervated myofibers. The antigen cannot be labeled in intact, whole muscle, but only in whole muscle that has been permeabilized with nonionic detergents. The antibody staining pattern in Triton X-100-permeabilized whole-mounts of the frog neuromuscular synapse is arranged in elongate, arborized areas which are characteristic of the frog neuromuscular synapse. The stained areas are striated and the striations occur with a periodicity that corresponds to the regular folding of the postsynaptic membrane. Immunoferritin labeling of fixed, saponin-permeabilized muscle demonstrates that the antigen is associated with amorphous material that is situated between the postsynaptic membrane and an underlying layer of intermediate filaments. The antigen, solubilized from membrane and an underlying layer of intermediate filaments. The antigen, solubilized from Torpedo electric organ by high ionic strength, was identified by antibody binding to nitrocellulose replicas of SDS gels of Torpedo tissue. In Torpedo tissue, the antibody binds to a single protein band at 51,000 daltons (51 kd). The 51-kd protein shares an antigenic determinant with intermediate filament proteins, since a monoclonal antibody to all intermediate filaments reacts with the same 51-kd protein. The monoclonal antibody also reacts with a 55-kd protein in frog skin which is localized to the perinuclear region of the epithelial cells.

Isolation and characterization of the mouse acetylcholine receptor delta subunit gene: identification of a 148-bp cis-acting region that confers myotube-specific expression.

We have isolated the gene encoding the delta subunit of the mouse skeletal muscle acetylcholine receptor (AChR) and have identified a 148-bp cis-acting region that controls cell type-specific and differentiation-dependent gene expression. The 5' flanking region of the delta subunit gene was fused to the protein-coding region of the chloramphenicol acetyltransferase (CAT) gene and gene fusions were transfected into C2 mouse skeletal muscle cells. Both transiently and stably transfected cells were assayed for CAT gene expression. Deletions from the 5' end of the mouse delta gene demonstrate that 148 bp of 5' flanking DNA is sufficient to confer cell type-specific and differentiation-dependent expression: CAT activity is present in transfected myotubes, but not in transfected 3T3 cells or 10T1/2 cells. Moreover, the level of CAT expression in myotubes transfected with constructs containing 148 bp of 5' flanking DNA from the delta subunit gene is identical to that in myotubes transfected with constructs containing 3.2 kb of 5' flanking DNA and similar to expression from the SV-40 early promoter. Increased CAT activity in myotubes is a result of an increased rate of transcription from the delta subunit promoter, since CAT RNA levels are also 35-fold more abundant in myotubes than myoblasts. In contrast, the SV-40 early promoter is similarly active in all cell types. Thus, 148 bp of 5' flanking DNA from the delta subunit gene contains all the information required for cell type-specific and differentiation-dependent expression of the AChR delta subunit.

300-kD subsynaptic protein copurifies with acetylcholine receptor-rich membranes and is concentrated at neuromuscular synapses.

Acetylcholine receptor-rich membranes from the electric organ of Torpedo californica are enriched in the four different subunits of the acetylcholine receptor and in two peripheral membrane proteins at 43 and 300 kD. We produced monoclonal antibodies against the 300-kD protein and have used these antibodies to determine the location of the protein, both in the electric organ and in skeletal muscle. Antibodies to the 300-kD protein were characterized by Western blots, binding assays to isolated membranes, and immunofluorescence on tissue. In Torpedo electric organ, antibodies to the 300-kD protein stain only the innervated face of the electrocytes. The 300-kD protein is on the intracellular surface of the postsynaptic membrane, since antibodies to the 300-kD protein bind more efficiently to saponin-permeabilized, right side out membranes than to intact membranes. Some antibodies against the Torpedo 300-kD protein cross-react with amphibian and mammalian neuromuscular synapses, and the cross-reacting protein is also highly concentrated on the intracellular surface of the post-synaptic membrane.

Acetylcholine receptors in regenerating muscle accumulate at original synaptic sites in the absence of the nerve.

We examined the role of nerve terminals in organizing acetylcholine receptors on regenerating skeletal-muscle fibers. When muscle fibers are damaged, they degenerate and are phagocytized, but their basal lamina sheaths survive. New myofibers form within the original basal lamina sheaths, and they become innervated precisely at the original synaptic sites on the sheaths. After denervating and damaging muscle, we allowed myofibers to regenerate but deliberately prevented reinnervation. The distribution of acetylcholine receptors on regenerating myofibers was determined by histological methods, using [125I] alpha-bungarotoxin or horseradish peroxidase-alpha-bungarotoxin; original synaptic sites on the basal lamina sheaths were marked by cholinesterase stain. By one month after damage to the muscle, the new myofibers have accumulations of acetylcholine receptors that are selectively localized to the original synaptic sites. The density of the receptors at these sites is the same as at normal neuromuscular junctions. Folds in the myofiber surface resembling junctional folds at normal neuromuscular junctions also occur at original synaptic sites in the absence of nerve terminals. Our results demonstrate that the biochemical and structural organization of the subsynaptic membrane in regenerating muscle is directed by structures that remain at synaptic sites after removal of the nerve.

Regulation of acetylcholine receptor transcript expression during development in Xenopus laevis.

The level of transcripts encoding the skeletal muscle acetylcholine receptor (AChR) was determined during embryonic development in Xenopus laevis. cDNAs encoding the alpha, gamma, and delta subunits of the Xenopus AChR were isolated from Xenopus embryo cDNA libraries using Torpedo AChR cDNAs as probes. The Xenopus AChR cDNAs have greater than 60% amino acid sequence homology to their Torpedo homologues and hybridize to transcripts that are restricted to the somites of developing embryos. Northern blot analysis demonstrates that a 2.3-kb transcript hybridizes to the alpha subunit cDNA, a 2.4-kb transcript hybridizes to the gamma subunit cDNA, and that two transcripts, of 1.9 and 2.5 kb, hybridize to the delta subunit cDNA. RNase protection assays demonstrate that transcripts encoding alpha, gamma, and delta subunits are coordinately expressed at late gastrula and that the amount of each transcript increases in parallel with muscle-specific actin mRNA during the ensuing 12 h. After the onset of muscle activity the level of actin mRNA per somite remains relatively constant, whereas the level of alpha subunit and delta subunit transcripts decrease fourfold per somite and the level of gamma subunit transcript decreases greater than 50-fold per somite. The decrease in amount of AChR transcripts per somite, however, occurs when embryos are paralyzed with local anaesthetic during their development. These results demonstrate that AChR transcripts in Xenopus are initially expressed coordinately, but that gamma subunit transcript levels are regulated differently than alpha and delta at later stages. Moreover, these results demonstrate that AChR transcript levels in Xenopus myotomal muscle cells are not responsive to electrical activity and suggest that AChR transcript levels are influenced by other regulatory controls.