Dietitians' Attitudes and Experiences of Blended Tube Feeding in Paediatrics

Aims To examine; (a) the number of registered dietitians (RDs) with blended tube fed (BTF) patients, work setting and caseload; (b) attitudes and experiences towards BTF; (c) current BTF supports and future resources required. Methods An online survey collected information from Irish RDs over one month. Data was examined using cross-tabulations and Mann-Whitney U tests. Free-text was categorized into thematic domains. Results A significant number of RDs with HEN paediatric patients concurrently managed BTF patients (n = 27/48, 56.3%, p<0.05). The majority were based in tertiary hospitals (HEN; n = 20/48, 41.7%, BTF; n = 12/27, 44.4%). Equal numbers were willing to support BTF or on a patient-case basis (n = 36/77, 46.8%). International guidelines were most used to inform RDs (n = 40/69, 58.0%). Professional training workshops were the preferred learning method (n = 60/73, 82.2%). Conclusion Overall, BTF appears to be a growing practice. Community services, professional guidelines, training and information are needed.

Outcomes of exclusive enteral nutrition in paediatric Crohn's disease.

BACKGROUND/OBJECTIVES: Exclusive enteral nutrition (EEN) is a safe and effective treatment modality for inducing remission in paediatric Crohn's disease (CD). The primary aim of this study was to compare the outcomes of EEN to corticosteroid (CS) therapy in newly diagnosed, treatment-naïve patients with CD. A secondary aim was to describe the outcomes of EEN in a national cohort of paediatric CD patients over a 10-year period. SUBJECTS/METHODS: A retrospective chart review was conducted at the Irish national referral centre for paediatric CD. A case-matched analysis was conducted on two cohorts matched for age, gender, disease location, disease behaviour and disease activity, who received CS or EEN as their initial treatment. Subsequently, cohort analysis was conducted on all patients who undertook a course of EEN therapy between 2004 and 2013. RESULTS: The case-matched analysis found higher remission rates after treatment with EEN (24/28, 86%) compared with those with CS (15/28, 54%; P=0.02). Dietetic contacts were found to be pivotal to the success of treatment and the attainment of remission. In total, 59 patients completed EEN at some time-point in their disease course and were included in the cohort analysis. Sixty-nine per cent of this cohort entered clinical remission (41/59). EEN was found to be most effective when used as an initial treatment (P=0.004) and less effective in patients aged under 10 years (P=0.04). CONCLUSIONS: EEN should be strongly considered as a favourable primary treatment over CS, especially in those diagnosed over the age of 10 years.

Sustained benefits of a community dietetics intervention designed to improve oral nutritional supplement prescribing practices.

BACKGROUND: Healthcare professionals working in the community do not always prescribe oral nutritional supplements (ONS) according to best practice guidelines for the management of malnutrition. The present study aimed to determine the impact of a community dietetics intervention on ONS prescribing practices and expenditure 1 year later. METHODS: The intervention involved general practitioners (GPs), practice nurses, nurses in local nursing homes and community nurses. It comprised an education programme together with the provision of a new community dietetics service. Changes in health care professionals' nutrition care practices were determined by examining community dietetics records. ONS prescribing volume and expenditure on ONS were assessed using data from the Primary Care Reimbursement Service of the Irish Health Service Executive. RESULTS: Seven out of 10 principal GPs participated in the nutrition education programme. One year later, screening for malnutrition risk was better, dietary advice was provided more often, referral to the community dietetics service improved and ONS were prescribed for a greater proportion of patients at 'high risk' of malnutrition than before (88% versus 37%; P < 0.001). There was a trend towards fewer patients being prescribed ONS (18% reduction; P = 0.074) and there was no significant change in expenditure on ONS by participating GPs (3% reduction; P = 0.499), despite a 28% increase nationally by GPs on ONS. CONCLUSIONS: The community dietetics intervention improved ONS prescribing practices by GPs and nurses, in accordance with best practice guidelines, without increasing expenditure on ONS during the year after intervention.

An evaluation of a community dietetics intervention on the management of malnutrition for healthcare professionals.

BACKGROUND: Healthcare professionals working in the community setting have limited knowledge of the evidence-based management of malnutrition. The present study aimed to evaluate a community dietetics intervention, which included an education programme for healthcare professionals in conjunction with the introduction of a community dietetics service for patients 'at risk' of malnutrition. Changes in nutritional knowledge and the reported management of malnourished patients were investigated and the acceptability of the intervention was explored. METHODS: An education programme, incorporating 'Malnutrition Universal Screening Tool (MUST)' training, was implemented in eight of 10 eligible primary care practices (14 general practitioners and nine practice nurses attended), in seven private nursing homes (20 staff nurses attended) and two health centres (53 community nurses attended) in conjunction with a community dietetics service for patients at risk of malnutrition. Nutritional knowledge was assessed before, immediately after, and 6 months after the intervention using self-administered, multiple-choice questionnaires. Reported changes in practice and the acceptability of the education programme were considered using self-administered questionnaires 6 months after the intervention. RESULTS: A significant increase in nutritional knowledge 6 months after the intervention was observed (P < 0.001). The management of malnutrition was reported to be improved, with 69% (38/55) of healthcare professionals reporting to weigh patients 'more frequently', whereas 80% (43/54) reported giving dietary advice to prevent or treat malnutrition. Eighty-percent (44/55) of healthcare professionals stated that 'MUST' was an acceptable nutrition screening tool. CONCLUSION: An education programme supported by a community dietetics service for patients 'at risk' of malnutrition increased the nutritional knowledge and improved the reported management of malnourished patients in the community by healthcare professionals.

The use of oral nutritional supplements in an Irish community setting.

BACKGROUND: The frequency of oral nutritional supplement (ONS) prescribing has been increasing steadily in the Republic of Ireland (ROI). Available evidence indicates that health professionals in the community setting in the ROI have a poor level of knowledge about ONS. The objectives of the present study were to investigate ONS prescribing practices and to identify the types of patient who were prescribed these products. METHODS: Ten of 17 eligible general practitioners were recruited and asked to refer all patients (aged > 16 years) who were prescribed ONS during a 3-month period. Patients were interviewed by a community dietitian, using a questionnaire incorporating the Malnutrition Universal Screening Tool (MUST). ONS prescriptions were judged either to fulfil or not to fulfil a set of criteria developed for ONS prescribing in the community. RESULTS: The majority of patients were female (62/78). Their mean (SD) age was 79 (10.5) years. According to MUST criteria, 31 of 78 patients were at 'low risk', 18 of 78 were at 'medium risk' and 29 of 78 were at 'high risk' of malnutrition. Less than half of the patients (36/78) had a body mass index of < 20 kg m(-2). Only 21 of 78 patients reported having received dietary advice in addition to their ONS prescription. Almost one-third (31%) of ONS prescriptions did not fulfil the criteria. Social factors, such as living alone, and difficulties with cooking and shopping, influenced the need for ONS in almost 70% of cases. CONCLUSIONS: ONS were prescribed in accordance with the prescribing criteria in the majority of cases; however, some patients who were prescribed ONS were not 'at risk' of malnutrition. Social circumstances played an important part in determining the need for ONS prescriptions.

Change in gene expression subsequent to induction of Pnn/DRS/memA: increase in p21(cip1/waf1).

Pnn (PNN) is a nuclear and cell adhesion-related protein. Previous work has suggested that Pnn/DRS/memA is a potential tumor suppressor involved in the regulation of cell adhesion and cell migration. Using the ecdysone-inducible mammalian expression system, a stable inducible GFP-tagged human Pnn gene (PNNGFP) expressing 293 cell line was created (EcR293-PNNGFP). Cells induced to express PNNGFP not only exhibited increased cell-cell adhesion but also exhibited changes in cell growth and cell cycle progression. cDNA array analyses, together with real time PCR, revealed that the effects of exogenously expressed Pnn on cellular behavior may be linked to the regulation of the expression of specific subset genes. This subset includes cell cycle-related genes such as p21(cip1/waf1), CDK4, CPR2; cell migration and invasion regulatory genes such as RhoA, CDK5, TIMP-1, MMP-7, and EMMPRIN; and MIC-1. Concordant with previous observations of Pnn-induced phenotype changes, genes coding for epithelial associated processes and cell division controls were elevated, while those coding for increased cell motility and cellular reorganizations were downregulated. We utilized p21 promoter-luciferase reporter constructs and demonstrated that a marked stimulation of p21 promoter activity in 293 cells correlated with increased Pnn expression. Taken together, these data indicate that Pnn may participate in the regulation of gene expression, thereby, positively promoting cell-cell adhesion, and negatively affecting cell migration and cell proliferation.

An organizational climate intervention associated with increased handwashing and decreased nosocomial infections.

Handwashing practices are persistently suboptimal among healthcare professionals and are also stubbornly resistant to change. The purpose of this quasi-experimental intervention trial was to assess the impact of an intervention to change organizational culture on frequency of staff handwashing (as measured by counting devices inserted into soap dispensers on four critical care units) and nosocomial infections associated with methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE). All staff in one of two hospitals in the mid-Atlantic region received an intervention with multiple components designed to change organizational culture; the second hospital served as a comparison. Over a period of 8 months, 860,567 soap dispensings were recorded, with significant improvements in the study hospital after 6 months of follow-up. Rates of MRSA were not significantly different between the two hospitals, but rates of VRE were significantly reduced in the intervention hospital during implementation.

Role of cell adhesion-associated protein, pinin (DRS/memA), in corneal epithelial migration.

PURPOSE: To determine whether the cellular distribution of cell adhesion-associated protein, pinin, is altered during corneal epithelial migration in response to debridement wounding and to determine the effect of overexpression of pinin in cultured epithelial cells. METHODS: Corneas from guinea pig and embryonic (day 17) chickens were excised, wounded, and placed on organ-culture rafts. At time points from 0 to 24 hours, corneas were cryosectioned and subsequently analyzed by immunofluorescence or immunoelectron microscopy for the presence and distribution of pinin. Cultured epithelial cell line MDCK (Madin Darby canine kidney) confluent monolayers were wounded by scraping and examined by immunofluorescence for pinin and desmoplakin. MDCK cells were transfected with full-length pinin cDNA. After selection in Geneticin, clones of pinin-transfected cells were isolated. Monolayers of transfected cells were scrape-wounded and assayed for their ability to migrate. RESULTS: Within 2 hours after wounding, although morphologically identifiable desmosomes were present on migrating epithelial cells, the association of pinin to desmosomes was greatly reduced. Finally, after completion of wound closure, pinin returned to the corneal epithelial desmosome. Wounding of confluent epithelial monolayers (MDCK) in vitro demonstrated a very similar change in the distribution of pinin, whereas desmoplakin remained cell boundary-associated. Transfection of pinin into cultured epithelial cells resulted in an overexpression of pinin. Clones of cells expressing high levels of pinin exhibited marked reduction in their ability to migrate after wounding. CONCLUSIONS: Pinin is involved in corneal epithelium migration. The localization of pinin at or near the desmosome is correlated with the epithelial quiescence. The loss of pinin from the cell boundary correlates with the transition from quiescence to actively migrating. Overexpressing pinin in cultured epithelial cells affects epithelial homeostasis and, in turn, drives the epithelial cells to a hyperstable epithelial adhesive state and inhibits the transition from quiescence to migratory.

Dissection of protein linkage between keratins and pinin, a protein with dual location at desmosome-intermediate filament complex and in the nucleus.

Pinin is a cell adhesion-associated and nuclear protein that has been shown to localize in the vicinity of intermediate filament (IF) convergence upon the cytoplasmic face of the desmosomal plaque as well as in the nucleus. The localization of pinin to the desmosomes has been correlated with the reinforcement of intercellular adhesion and increased IF organization. In this study, keratins 18, 8, and 19 were identified to interact with the amino end domain of pinin in a two-hybrid screening. Further truncation analyses indicated that the 2B domain of keratin contains the sequence responsible for interacting with pinin. The amino end of pinin (residues 1-98) is sufficient to bind to keratin. Point mutation analyses revealed two essential residues within the pinin fragment 1-98, leucine 8 and leucine 19, for the interaction with keratin. Finally, in vitro protein overlay binding assays confirmed the direct interaction of the amino end domain of pinin with keratins, while pinin mutant L8P GST fusion protein failed to bind to keratins in the overlay assay. Coupled with our previous morphological observations and transfection studies, these data suggest that pinin may play a role in epithelial cell adhesion and the IF complex through a direct interaction with the keratin filaments.

Characterization of the gene encoding pinin/DRS/memA and evidence for its potential tumor suppressor function.

Several cell adhesion-related proteins have been shown to act as tumor-suppressors (TS) in the neoplastic progression of epithelial-derived tumors. Pinin/DRS/memA was first identified in our laboratory and it was shown to be a cell adhesion-related molecule. Our previous study demonstrated that restoration of pinin expression in transformed cells not only positively influenced cellular adhesive properties but also reversed the transformed phenotype to more epithelial-like. Here, we show by FISH analysis that the gene locus for pinin is within 14q13. The alignment of the pinin gene with STS markers localized the gene to the previously identified TS locus D14S75-D14S288. Northern analyses revealed diminished pinin mRNA in renal cell carcinomas (RCC) and certain cancer cell lines. Immunohistochemical examination of tumor samples demonstrated absent or greatly reduced pinin in transitional cell carcinoma (TCC) and RCC tumors. TCC-derived J82 cells as well as EcR-293 cells transfected with full-length pinin cDNA demonstrated inhibition of anchorage-independent growth of cells in soft agar. Furthermore, methylation analyses revealed that aberrant methylation of pinin CpG islands was correlated with decreased/absent pinin expression in a subset of tumor tissues. These data lend significant support to the hypothesis that pinin/DRS/memA may act as a tumor suppressor in certain types of cancers.

Dietary intakes in Ireland of a healthy elderly population.

The aim of this paper is to assess the adequacy of the diet of individuals over 60 yr of age, participating in the 1990 Irish National Nutritional Survey. A nationwide random sample based on the most recently updated electoral register was used. Demographic information was collected. Anthropometric measurements were taken and nutrient intake was assessed using the 7-day dietary history method. The randomly selected sample of 1213 subjects was considered to be representative of the Irish population. Of those selected, 163 individuals were over 60 yr of age, 79 of whom were male and 84 female. Mean energy intakes including alcohol for males and females were 9.55 +/- 3.09MJ and 7.07 +/- 2.39MJ respectively. The main sources of energy were bread, meat and meat products, potatoes and milk. As percentage energy, protein, fat and carbohydrate intakes were 14.90 per cent, 33.97 per cent and 48.22 per cent for men and 15.39 per cent, 34.09 per cent and 49.37 per cent for women respectively. Except for vitamin D and folate, micronutrient intakes were adequate. The body mass index (BMI [weight/height2] kg/m2) for men was 25.6 and for women 26.4. Fewer than 27.8 per cent of the males and 20.2 per cent of females take part in regular physical activity. In conclusion, the diet of a healthy elderly population in Ireland is nutritionally adequate with macronutrient intake in keeping with the recommended guidelines. Overall energy intakes are lower than those of a younger age group and may account for the lower intakes of certain micronutrients. An increase in fruit and vegetable consumption would improve vitamin and mineral intake. In order to allow for a higher energy intake an increase in physical activity is desirable.

Cloning and analysis of cDNA encoding murine pinin.

Pinin is a cell junction-associated protein involved in the stabilization of the desmosome-intermediate filament complex in various epithelial tissues. Utilizing a cDNA probe derived from canine pinin, we isolated overlapping cDNA clones encoding murine full-length pinin. The total cDNA contained an open reading frame of 2175 nucleotides coding for 725 amino acids as well as a 3'- and a 5'-untranslated regions of 620 and 18 nucleotides, respectively. The overall predicted amino acid sequence of mouse pinin displayed strong identities to those of canine and human pinin, with the exception of a stretch of 38 amino acids which were found to be deleted in mouse pinin. There were several discernible domains found within mouse pinin. These included three coiled-coil domains, a small stretch of glycine loops, a short glutamine-proline-rich domain and a polyserine domain.

ZO1 in corneal epithelium: association to the zonula occludens and adherens junctions.

The corneal epithelium, like other stratifying epithelium, does not present a well formed junctional complex as compared to that of simple epithelia. However, the resistance barrier of the corneal epithelium is to a great extent generated by zonula occludens (ZO, tight junction), which are formed between the cells of the apical-most strata. The tight junction provides a continuous seal around the apical aspect of adjoining epithelial cells, thereby preventing the free passage of molecules between adjacent epithelial cells (paracellular pathway). We have examined rabbit corneal epithelia with monoclonal antibody against the tight junction associated protein ZO1. With this antibody, we resolved two distinct patterns of ZO1 expression, one being the lateral boundary of the apical cell, which appeared as a true zonula around these cells. The second pattern of expression for ZO1 was at a set of punctate spots that correspond to the connection of the most apical portion of the basal corneal epithelial cells, with the above wing cells. En face, confocal analyses revealed that these areas consisted of 5-6 distinct spots per basal cell at or near the contact points with the immediate wing cells above. ImmunoEM revealed that the mid-epithelial accumulations of ZO1 were not tight junctions, but rather a form of adherens junction. The expression of ZO1 in the mid-epithelial level of the cornea is neither correlated with the presence of tight junction, nor with the established barrier functions. Interestingly, these junctions in the corneal epithelium also contain paxillin, a focal adhesion associated phosphoprotein which is a target of pp125 focal adhesion kinase, erbB-2 kinase and p21Obcr/abl oncogene. We postulated that the ZO1/paxillin adherens junction within stratified epithelium, such as the corneal epithelium, may function to reinforce attachments at the level of the basal cell to wing cell junction and be regulated by reversible phosphorylation. We speculate that the regulated phosphorylation of tyrosine residues on paxillin may perform a critical role in controlling epithelial cell-cell interactions as it does in cell-matrix adhesion.

A novel repressor, par-4, modulates transcription and growth suppression functions of the Wilms' tumor suppressor WT1.

The tumor suppressor WT1 represses and activates transcription. The loss and/or imbalance of the dual transcriptional activity of WT1 may contribute to Wilms' tumor. In this study, we identified par-4 (for prostate apoptosis response) as a WT1-interacting protein that itself functions as a transcriptional repressor. par-4 contains a putative leucine zipper domain and is specifically upregulated during apoptosis of prostate cells (S. F. Sells, D. P. Wood, Jr., S. S. Joshi-Barve, S. Muthukkumar, R. J. Jacob, S. A. Crist, S. Humphreys, and V. M. Rangnekar, Cell Growth Differ. 5:457-466, 1994). The leucine repeat domain of par-4 was shown to interact with the zinc finger DNA binding domain of WT1. Immunoprecipitation-Western blot (immunoblot) analyses demonstrated in vivo WT1-par-4 interactions. par-4 was ubiquitously expressed, and the protein was found in both the nucleus and the cytoplasm. Functionally, par-4 inhibited transcription activated by WT1, but not by the related protein EGR1. Inhibition of WT1-mediated transcription was dependent on the domain of par-4 that mediates its physical association with WT1. In addition, par-4 augmented WT1-mediated repression, possibly by contributing an additional repression domain. Consistent with these results, par-4 functioned as a transcriptional repressor when brought to a promoter via a heterologous DNA binding domain. Significantly, par-4, but not a mutant unable to interact with WT1, rescued growth suppression caused by WT1. Thus, we identified a novel repressor that modulates transcription as well as growth suppression functions of WT1.

Characterization of pinin, a novel protein associated with the desmosome-intermediate filament complex.

We have identified a protein named pinin that is associated with the mature desmosomes of the epithelia (Ouyang, P., and S.P. Sugrue. 1992. J. Cell Biol. 118:1477-1488). We suggest that the function of pinin is to pin intermediate filaments to the desmosome. Therefore, pinin may play a significant role in reinforcing the intermediate filament-desmosome complex. cDNA clones coding for pinin were identified, using degenerative oligonucleotide probes that were based on the internal amino acid sequence of pinin for the screening of a cDNA library. Immunoblotting of expressed recombinant proteins with the monoclonal 08L antibody localized the 08L epitope to the carboxyl end of the protein. Polyclonal antibodies directed against fusion proteins immunoidentified the 140-kD protein in tissue extracts. Immunofluorescence analysis, using the antifusion protein antibody, demonstrated pinin at lateral epithelial boundaries, which is consistent with desmosomal localization. The conceptual translation product of the cDNA clones contained three unique domains: (a) a serine-rich domain; (b) a glutamine-proline, glutamine-leucine repeat domain; and (c) an acidic domain rich in glutamic acid. Although the 3' end of the open reading frame of the clone for pinin showed near identity to a partial cDNA isolated for a pig neutrophil phosphoprotein (Bellavite, P., F. Bazzoni, et al. 1990. Biochem. Biophys. Res. Commun. 170:915-922), the remaining sequence demonstrated little homology to known protein sequences. Northern blots of mRNA from chicken corneal epithelium, MDCK cells, and various human tissues indicated that pinin messages exhibit tissue-specific variation in size, ranging from 3.2 to 4.1 kb. Genomic Southern blots revealed the existence of one gene for pinin, suggesting alternative splicing of the mRNA. Expression of the full-length cDNA clones in human 293 cells and monkey COS-7 cells demonstrated that a 140-kD immunoreactive species on Western blots corresponded to pinin. Pinin cDNA transfected into the transformed 293 cells resulted in enhanced cell-cell adhesion. Immunofluorescence staining revealed that the expressed pinin protein was assembled to the lateral boundaries of the cells in contact, which is consistent with the staining pattern of pinin in epithelial cells.

Hematopoietic supportive functions of mouse bone marrow and fetal liver microenvironment: dissection of granulocyte, B-lymphocyte, and hematopoietic progenitor support at the stroma cell clone level.

We dissected the functions of the microenvironment of bone marrow (BM) and fetal liver (FL) at the cellular level by cloning individual stromal calls and characterizing their phenotypical and functional features. Stromal cell clones derived from FL are large in size (mean forward light scatter intensity [mFSC] of 450), express the surface antigen Thy-1 but not Sca-1 and 6 out of 6 are able to differentiate into fat accumulating adipocytes. BM derived stromal cell clones are either small (mFSC of 250) or large (mFSC of 450), express Sca-1 but not Thy-1 and only 2 out of 7 differentiate towards adipocytes. Heterogeneity in terms of vascular adhesion molecule-1, intracellular adhesion molecule-1 and heat stable antigen expression was found among the different cell clones. Functional assays using long- and short-term cocultures of stromal and hematopoietic calls revealed: (1) the capacity of 8 out of 12 stromal cell clones to support the expansion of primitive hematopoietic progenitors (colony forming unit spleen day 12) more than 10 weeks. Fat accumulation but not expression of stem cell factor by stromal cells did correlate with this supportive function. (2) Better support of granulocyte maturation and proliferation by BM- compared to FL-derived stromal cell clones. However, stromal cell clones from both organs expressed macrophage-colony stimulating factor. (3) The ability of 4 out of 12 stromal cell clones (derived from both, FL and BM) to support the expansion of Interleukin-7 dependent pre-B cells from the BM. Pre-B cell growth stimulating factor was not restricted to supporters. (4) Mutual exclusiveness of myeloid and lymphoid support in that a given stromal cell clone supported either pre B-cell or granulocyte expansion. Experiments comparing the support of BM- and FL-derived hematopoietic progenitors showed identical responses of late (B220+/c-kit-) but strikingly different responses of early (B220+/c-kit+) pre-B cells, revealing different proliferation requirements for FL- versus BM- derived early pre-B cells in vitro.

Histopathologic features of the floppy eyelid syndrome. Involvement of tarsal elastin.

PURPOSE: Patients with the floppy eyelid syndrome have chronic papillary conjunctivitis with easily everted upper eyelids and a soft, pliant upper tarsus. The purpose of this study is to describe the clinical features and the histopathologic correlate in a group of patients with floppy eyelid syndrome. METHODS: The authors examined eight patients with floppy eyelid syndrome, four of whom underwent surgical management with horizontal eyelid shortening. Eyelid tissue from these patients was examined using light microscopy, electron microscopy, and immunohistochemistry and compared with controls with unrelated eyelid or orbital disorders. RESULTS: Clinical findings included obesity or eye rubbing, lash ptosis, and, less commonly, blepharoptosis. Two patients had documented sleep apnea with abnormal sleep electroencephalogram. Light microscopy of the surgical specimens showed chronic conjunctival inflammation, papillary conjunctivitis, and meibomian gland abnormalities, including granuloma formation. Verhoeff's modified elastin stain demonstrated a marked decrease in the amount of elastin fibers in tarsus from patients with floppy eyelid syndrome compared with controls. Immunohistochemical staining for elastin also showed a marked decrease of tarsal elastin in floppy eyelid patients compared with controls. In contrast, immunohistochemical stains showed that the distribution of collagen types I and III was similar between patients with floppy eyelid syndrome and controls. Electron microscopy demonstrated that tarsal collagen was comparable in patients and controls, and that there was a reduced amount of tarsal elastin in floppy eyelid syndrome compared with controls. CONCLUSIONS: These findings demonstrate that tarsal elastin is decreased in the floppy eyelid syndrome, which may contribute to the laxity of the tarsus in this disorder.

Identification of an epithelial protein related to the desmosome and intermediate filament network.

Using a mAb, referred to as 08L, we have identified a protein, of M(r) approximately 140,000, associated with desmosomes of epithelial cells. The 08L antibody stained the intracellular side of lateral cell margins of monolayer epithelial cells but did not stain cell margins free of cell contact. Immunoelectron microscopy revealed that the 08L antigen was localized to the cytosolic surface of the desmosomal plaque near points of intermediate filament convergence with apparently little staining of the desmosomal plaque proper. Western blots revealed the 08L antigen to be a protein, of M(r) approximately 140,000, found in the Triton-X 100 insoluble pellet. High salt-containing buffers extracted the 08L antigen from the insoluble material. Examination of the assembly of 08L to the desmosome complex, in cells grown in low confluent culture or in calcium-switch assays, by double immunofluorescence with 08L and anti-desmoplakin antibody, revealed that 08L was recruited to morphologically identifiable desmosomes. 08L antigen may exist in a cytosolic pool prior to assembly to the cell surface. The solubility of 08L in low calcium and normal calcium conditions, however, was similar. 08L association to the desmosome was correlated with increased organization of the intermediate filament network. We suggest that the 08L antigen may be involved in the organization and stabilization of the desmosome-IF complexes of epithelia.

Immunolocalization of type XII collagen at the corneoscleral angle of the embryonic avian eye.

A monoclonal antibody specific for the chicken alpha 1 (XII) collagen chain was used to immunolocalize type XII collagen in the corneoscleral of 17-19-day-old chicken embryos. These immunofluorescence studies localized type XII collagen around the scleral cartilages and ossicles. There was also a striking positive staining in the scleral spur and stroma of the corneolimbus beneath the external scleral sulcus. By immunofluorescence the corneal stroma proper was found to be relatively poor in type XII collagen. Immunoblot analysis confirmed the relative absence of alpha 1(XII) in the corneal stroma and demonstrated the presence of a 240 -kD polypeptide representative of alpha 1(XII) in extracts of the corneoscleral angle. It was suggested that type XII collagen in the corneoscleral angle reinforces the collagenous matrix at specific sites which require high tensile strength.