The volume of liver irradiated during modern free-breathing breast radiotherapy: Implications for theory and practice.

INTRODUCTION: Incidental liver irradiation during breast radiotherapy can increase the risk of second primary malignancy and induce adverse inflammatory states. This study establishes the volume of liver irradiated during free-breathing breast radiotherapy. Novel associations between liver dose-volume data and systemic interleukin-6 soluble receptor and blood counts are evaluated. METHODS: The volume of liver within the 10%, 50% and 90% isodose was determined for 100 women with stage 0 to II breast carcinoma undergoing 40Gy in 15 fractions over three weeks tangential irradiation. Blood counts and interleukin 6 soluble receptor concentration were recorded before, during and four weeks after radiotherapy. Dose-volume data for right-sided treatments was associated with longitudinal measures at bivariate and multivariable levels. RESULTS: A maximum of 226cm3 (19%), 92 cm3 (8%) and 62 cm3 (5%) of the liver was irradiated within the 10%, 50% and 90% isodose. Liver irradiation was almost exclusively a feature of the 52 right-sided treatments and was strongly correlated with breast volume (ρ = 0.7, p < 0.0001). Liver V10% was significantly associated with interleukin-6 soluble receptor concentration four weeks post-radiotherapy (beta = 0.38, p = 0.01) after controlling for theoretical confounding variables. CONCLUSION: Up to 8% of the liver is irradiated within the primary beam during local right-sided breast radiotherapy. Select use of a deep inspiration breath hold technique would reduce this volume, and minimise the risk of radiation-induced malignancy and acute systemic elevation of inflammatory interleukin 6 soluble receptor.

ACTIVE - a randomised feasibility trial study protocol of a behavioural intervention to reduce fatigue in women undergoing radiotherapy for early breast cancer: study protocol.

BACKGROUND: Fatigue is rated as the most distressing side effect of radiotherapy treatment for curable breast cancer. About four in ten women treated experience fatigue, which can last for years after treatment. The impact of this debilitating tiredness is loss of independence and impaired physical and mental function. Our study will take a behavioural intervention with demonstrated effect in treating fatigue in a mixed group of chemotherapy patients and adapt it for women undergoing radiotherapy for early breast cancer. The purpose of this trial is to evaluate the feasibility of delivering the intervention in the radiotherapy pathway for patients at a high risk of fatigue and to explore participants' experiences of the trial and intervention. METHODS: A pragmatic single-site non-blinded feasibility trial of a behavioural intervention. Main inclusion criteria are prescription of the UK standard 40 Gy in 15 fractions over 3 weeks of radiotherapy (± tumour bed boost) for early (stage 0-IIIa) breast cancer. The total projected sample size after attrition is 70. A previously developed fatigue risk score tool will be used to predict individual's likelihood of experiencing fatigue. Thirty women predicted to be at a high risk of experiencing significant fatigue will be allocated in the ratio 2:1 to the behavioural intervention or education trial arms, respectively. These feasibility trial participants will be assessed at baseline, after 10 and 15 fractions of radiotherapy and 10 days, 3 weeks and 6 months after radiotherapy. A further 40 women predicted to be at a lower risk of fatigue will join a risk score validation group.Measures to assess feasibility include recruitment, retention and completion rates and variation in implementation of the intervention. Process evaluation with intervention providers and users includes fidelity and adherence checks and qualitative interviews to understand how changes in behaviour are initiated and sustained. DISCUSSION: This feasibility study collates data to both inform the progression to and design of a future definitive trial and to refine the intervention. TRIAL REGISTRATION: ISRCTN 10303368. Registered August 2017 (retrospectively registered); Health and Care Research Wales Clinical Research Portfolio (CRP) registration 31419.

A prognostic tool to predict fatigue in women with early-stage breast cancer undergoing radiotherapy.

BACKGROUND: Fatigue during and after radiotherapy impacts negatively on normal functioning and quality of life. A pre-treatment estimate of the risk of fatigue would facilitate the targeting of timely interventions to limit consequential behavioural symptoms arising. We have developed a prognostic tool to predict the risk of fatigue in women with early-stage breast cancer undergoing radiotherapy. METHODS: Socio-demographic, clinical and self-reported characteristics were recorded for 100 women prescribed adjuvant radiotherapy for stages Tis-T2N1 breast cancer. Multiple logistic regression was used to develop a parsimonious prognostic model. The performance of the model when predicting fatigue for individuals not in the study was estimated by a leave-one-out cross-validation. A statistical weighting was assigned to the model variables to render a Fatigue Propensity Score of between 0 and 15. The ability of the Propensity Score to discriminate fatigued participants was estimated via receiver operating characteristic curve analysis. RESULTS: 38% of participants reported significant fatigue during radiotherapy. Fatigue risk was predicted by elevated pre-treatment fatigue and anxiety, and diagnoses other than invasive ductal carcinoma (ductal carcinoma in-situ, invasive lobular and rarer carcinoma subtypes). The positive predictive value of the prognostic model was 80%. A Propensity Score threshold of ≥6 corresponded to a specificity of 90.3% and a sensitivity of 76.3%. The area under the receiver operating characteristic curve was 0.83 for the cross-validation sample. CONCLUSIONS: Application of the Fatigue Propensity Score in the patient pathway can help direct fatigue management resources at those patients most likely to benefit.

Physicochemical basis for dilated intercellular spaces in non-erosive acid-damaged rabbit esophageal epithelium.

Dilated intercellular spaces (DIS) within esophageal epithelium (EE) is a histopathologic feature of non-erosive reflux disease and early lesion in acid-damaged rabbit EE associated with increased paracellular permeability. Its cause remains unknown, but the lesion's morphology suggests a significant fluid shift into the intercellular spaces (ICS). Since water follows osmotic forces and consequently ion movements, we explored the role of active (ion) transport and ion gradients in its pathogenesis. This was done by quantifying the effect of inhibited active transport and altered ion gradients on electrical resistance (R(T)) and ICS diameter in acid-exposed Ussing-chambered rabbit EE. Compared with normal Ringer, pH 7.5, 30 minutes of luminal HCl (100 mmol/L), pH 1.1, increased permeability (R(T): +5 +/- 4% vs-52 +/- 4%) and ICS diameter (0.25 +/- 0.01 microm vs 0.42 +/- 0.02 microm), but had no effect on cell morphology or diameter. Ouabain pretreatment significantly reduced active transport but had no effect on the acid-induced changes. However, negating the chloride gradient created by luminal HCl either by adding choline chloride, 100 mmol/L, serosally or by replacing luminal HCl, pH 1.1, with luminal H(2)SO(4), pH 1.1, prevented the development of DIS while maintaining the increase in permeability. DIS was also prevented in the presence of a 100 mmol/L (choline) chloride gradient by luminal exposure at neutral pH. DIS in HCl-damaged EE is caused by an H(+)-induced increase in epithelial permeability; this enables Cl(-) to diffuse along its gradient into the ICS, creating an osmotic force for water movement into and (hydrostatic) dilation of the ICS.

Freeze fracture study of airway epithelium from patients with primary ciliary dyskinesia.

BACKGROUND: The airway cilia of patients with primary ciliary dyskinesia (PCD) exhibit several anomalies when studied by transmission electron microscopy, but little is known about the ultrastructural organisation of ciliary membranes in these patients. Freeze fracture replication of airway epithelium from patients with PCD provides a means of achieving high resolution views of cell membrane structure. Ciliary necklaces are a specialised structural feature of ciliary membranes thought to serve as a timing mechanism for ciliary beat, and their characterisation in the cilia of patients with PCD may contribute new insights into the pathophysiology of this syndrome. METHODS: The nasal epithelium of three patients with PCD was freeze fractured and replicated with platinum and carbon shadowing. The resultant preparations were examined by transmission electron microscopy and the ciliary necklaces were compared with similar preparations of nasal biopsy specimens from normal healthy subjects. RESULTS: The ciliary necklaces of the three patients with PCD were normal with no overt differences from those of healthy individuals. CONCLUSIONS: The defective ciliary motility observed in patients with PCD does not appear to result from membrane dysfunction associated with overt disorganisation of ciliary necklace structure.

Canines as sentinel species for assessing chronic exposures to air pollutants: part 1. Respiratory pathology.

A complex mixture of air pollutants is present in the ambient air in urban areas. People, animals, and vegetation are chronically and sequentially exposed to outdoor pollutants. The objective of this first of 2 studies is to evaluate by light and electron microscopy the lungs of Mexico City dogs and compare the results to those of 3 less polluted cities in MEXICO: One hundred fifty-two clinically healthy stray mongrel dogs (91 males/61 females), including 43 dogs from 3 less polluted cities, and 109 from southwest and northeast metropolitian Mexico City (SWMMC, NEMMC) were studied. Lungs of dogs living in Mexico City and Cuernavaca exhibited patchy chronic mononuclear cell infiltrates along with macrophages loaded with particulate matter (PM) surrounding the bronchiolar walls and extending into adjacent vascular structures; bronchiolar epithelial and smooth muscle hyperplasia, peribronchiolar fibrosis, microthrombi, and capillary and venule polymorphonuclear leukocytes (PMN) margination. Ultrafine PM was seen in alveolar type I and II cells, endothelial cells, interstitial macrophages (Mtheta), and intravascular Mtheta-like cells. Bronchoalveolar lavage showed significant numbers of alveolar macrophages undergoing proliferation. Exposure to complex mixtures of pollutants-predominantly particulate matter and ozone-is causing lung structural changes induced by the sustained inflammatory process and resulting in airway and vascular remodeling and altered repair. Cytokines released from both, circulating inflammatory and resident lung cells in response to endothelial and epithelial injury may be playing a role in the pathology described here. Deep concern exists for the potential of an increasing rise in lung diseases in child populations exposed to Mexico City's environment.

Canines as sentinel species for assessing chronic exposures to air pollutants: part 2. Cardiac pathology.

The principal objective of this study is to evaluate by light and electron microscopy (LM, EM) the heart tissues in stray southwest and northeast metropolitan Mexico City (SWMMC, NEMMC) dogs and compare their findings to those from 3 less polluted cities (Cuernavaca, Tlaxcala, and Tuxpam). Clinically healthy mongrel dogs, including 109 from highly polluted SWMMC and NEMMC, and 43 dogs from less polluted cities were studied. Dogs residing in cities with lower levels of pollutants showed little or no cardiac abnormalities. Mexico City and Cuernavaca dogs exhibited LM myocardial alterations including apoptotic myocytes, endothelial and immune effector cells, degranulated mast cells associated with scattered foci of mononuclear cells in left and right ventricles and interventricular septum, and clusters of adipocytes interspersed with mononuclear cells. Vascular changes included scattered polymorphonuclear leukocytes (PMN) margination and microthrombi in capillaries, and small venous and arteriolar blood vessels. Small veins exhibited smooth muscle cell hyperplasia, and arteriolar blood vessels showed deposition of particulate matter (PM) in the media and adventitia. Unmyelinated nerve fibers showed endoneural and epineural degranulated mast cells. EM examination of myocardial mast cells showed distended and abundant rough endoplasmic reticulum with few secretory granules. Myocardial capillaries exhibited fibrin deposition and their endothelial cells displayed increased luminal and abluminal pinocytic activity and the formation of anemone-like protrusions of the endothelium into the lumen. A close association between myocardial findings, lung epithelial and endothelial pathology, and chronic inflammatory lung changes was noted. The myocardial changes described in dogs exposed to ambient air pollutants may form the basis for developing hypothesis-driven mechanistic studies that might explain the epidemiological data of increased cardiovascular morbidity and mortality in people exposed to air pollutants.

Ultrastructural nasal pathology in children chronically and sequentially exposed to air pollutants.

Southwest Metropolitan Mexico City (SWMMC) children are repeatedly exposed to a complex mixture of air pollutants, including ozone, particulate matter, and aldehydes. Nasal biopsies taken from these children exhibit a wide range of histopathologic alterations: marked changes in ciliated and goblet cell populations, basal cell hyperplasia, squamous metaplasia, and mild dysplasias. We studied the ultrastructural features of 15 nasal biopsies obtained from clinically healthy children 4 to 15 yr of age, growing up in SWMMC. The results were compared with nasal biopsies from 11 children growing up in Veracruz and exposed to low pollutant levels. Ultrathin sections of nasal biopsies revealed an unremarkable mucociliary epithelium in control children, whereas SWMMC children showed an epithelium comprised of variable numbers of basal, ciliated, goblet, and squamous metaplastic as well as intermediate cells. Nascent ciliated cells, as evidenced by the presence of migratory kinetosomes, were common, as were ciliary abnormalities, including absent central microtubules, supernumerary central and peripheral tubules, ciliary microtubular discontinuities, and compound cilia. Dyskinesia associated with these abnormal cilia was suggested by the altered orientation of the central microtubules in closely adjacent cilia. A transudate was evident between epithelial cells, suggesting potential deficiencies in epithelial junction integrity. Particulate matter was present in heterolysosomal bodies in epithelial cells and it was also deposited in intercellular spaces. The severe structural alteration of the nasal epithelium together with the prominent acquired ciliary defects are likely the result of chronic airway injury in which ozone, particulate matter, and aldehydes are thought to play a crucial role. The nasal epithelium in SWMMC children is fundamentally disordered, and their mucociliary defense mechanisms are no longer intact. A compromised nasal epithelium has less ability to protect the lower respiratory tract and may potentially leave the distal acinar airways more vulnerable to reactive gases. Impairment of mucociliary clearance has the potential to increase the contact time between deposited mutagenic particulate matter and the epithelial surface, thus increasing the risk for nasal carcinogenesis. Chronic exposures to air pollutants affect the whole respiratory tract; the nasal epithelium is an accessible and valuable sentinel to monitor exposures to toxic or carcinogenic substances.

Characterization of an axonemal dynein heavy chain expressed early in airway epithelial ciliogenesis.

The most conspicuous evidence of airway epithelial maturation and vitality is the presence of motile cilia. In an effort to generate genetic and antigenic markers of airway maturation, injury, and repair, we characterized airway epithelial expression of a gene identified by two human expressed sequence tags that encoded peptides with sequence similarity to an invertebrate ciliary dynein heavy chain (DHC). Molecular analyses showed that the gene has a very large RNA transcript that encodes a very high molecular weight polypeptide with biochemical properties that are characteristic of a dynein heavy chain. Expression of the gene transcript correlated with the presence of ciliated cells in tissues, and immunohistochemical localization of the gene product confirmed its presence in the cilia of mature airway epithelium. In epithelium undergoing ciliogenesis ex vivo, expression of the gene transcript preceded ciliation of the epithelium and the gene product was present in the cytoplasm and at the apical border of nonciliated cells. These data suggested that the gene encodes an axonemal DHC that is expressed early during ciliogenesis, before the appearance of cilia.

An autoradiographic assessment of epithelial cell proliferation and post-natal maturation of the tracheal epithelium in infant ferrets.

The tracheal epithelium of infant ferrets undergoes rapid postnatal maturation over the first month of life to achieve the pseudostratified columnar configuration characteristic of the large airways of other mammals. We have used in vivo pulsing with tritiated thymidine ((3)HT) to elicit autoradiographic labeling of cells synthesizing nucleic acids in order to characterize more fully the contribution to development of different cell types comprising the nascent epithelial layer during this period of rapid growth. These studies indicate that two distinct populations of epithelial cells possess proliferative potential and contribute to the establishment of the mature adult epithelial layer. These investigations further confirm the mitotic potential of basal cells during a period of rapid postnatal growth and development of the tracheal epithelial layer. These studies also document the contribution to early airway development by non-ciliated cells, which predominate on the luminal border of the ferret trachea at birth. The temporal and histologic patterns of airway epithelial maturation during post-natal life in the ferret as contained in this study exhibit similarities to those which occur with recovery from injury by infection and irritant exposure in mature airways. Thus, the characterization of epithelial cell compartments having proliferative potential may provide insights into the mechanisms whereby normal airway epithelial organization is established and maintained during development as well as the possible recapitulation of these mechanisms during times of epithelial regeneration following injury.

Connexin 26 expression in human and ferret airways and lung during development.

Coordinated microscopic and molecular biological studies were used to document gap junction expression during postnatal development in ferret tracheal epithelium and lung and in fetal and adult human airway and lung. Expression of connexin 26 (Cx26) in the ferret airways was limited to the epithelial layer and was observed only during the newborn interval. In contrast, we found Cx26 expressed in the alveolar epithelium of the ferret lung by in situ hybridization, Northern blotting, RT-PCR amplification, and immunocytochemical labeling at all ages examined. This finding was further confirmed by documentation of gap junctional plaques upon ultrastructural examination of freeze-fracture replicas of adult ferret lung tissue. Parallel studies of developing human fetal lung and airway suggested connexin expression in the airways only in the first trimester but, as in the ferret, persistent expression was observed in both fetal and adult lung. These studies suggest that the transient expression of Cx26 is a reliable early indicator of airway epithelial development and differentiation in the airways. In contrast, Cx26 expression persists throughout life in the lung, suggesting that gap junctions serve more perennial intercellular communication functions in the peripheral lung.

Bubble hair. Case caused by an overheating hair dryer and reproducibility in normal hair with heat.

BACKGROUND: Bubble hair is an acquired hair shaft deformity characterized by bubble-like areas in the hair shaft seen with light microscopy and corresponding cavitary defects with scanning electron microscopy. OBJECTIVE: Our purpose was to report the fourth case of bubble hair, to demonstrate a cause, and to determine whether the cavities contain gas or liquid. METHODS: Light and scanning electron microscopy were performed. The patient's hair dryer was examined. We applied heat to normal hair of the patient and that of 16 human volunteers. Approximate temperatures for bubble formation were measured. The bubble contents were investigated by applying aqueous and nonaqueous liquids to affected hairs. RESULTS: No person's hair failed to develop bubbles when subjected to sufficient heat. The cause of bubble hair in the patient was an overheating hair dryer. The cavitations contained a gas. CONCLUSION: Bubble hair is a reproducible hair shaft defect caused by heat. The use of overheating hair dryers, or any other hair care equipment that overheats, should be avoided.

Ultrastructure of airway epithelial cell membranes among patients with cystic fibrosis.

Alterations in transepithelial ion fluxes are a primary pathophysiologic feature in cystic fibrosis (CF). Chronic respiratory infections and host responses are secondary aggravating pulmonary complications of this disease. In the present study, the application of the freeze-fracture technique to samples of nasal and tracheal epithelium from patients with CF has provided a perspective of large areas of cell membrane for the evaluation of possible structural correlates to the pathophysiology of this disease. A variety of aberrant configurations in stranding pattern and disorganization of the epithelial tight junctional complexes in CF airway epithelium are described. Additionally, examination of ciliary membranes revealed the presence of compound cilia and dysmorphology of ciliary necklace configuration. These features are thought to represent acquired structural lesions possibly derived from chronic infection and/or host responses which may further exacerbate abnormal ion transport properties and decrements of ciliary function that appear to be associated with the airway epithelium of individuals with CF.

Dynamics of intercellular communication and differentiation in a rapidly developing mammalian airway epithelium.

A considerable body of data suggests that gap junctions represent channels that facilitate intercellular communication, thereby modulating growth and development. However, direct quantitative evidence supporting such a structure/function relationship is limited. This study has identified a new model in the rapidly developing tracheal epithelium of infant ferrets wherein gap junction prevalence and intercellular transfer of a fluorescent, low molecular weight dye, epithelial cell incorporation of tritiated thymidine, and progressive ciliation of the epithelium have been characterized. This developmental pattern provides favorable conditions for the study of relationships between gap junctions, intercellular translocation of chemical signals, and cell growth and differentiation in a mammalian airway epithelium with a minimum of experimental intervention.

Development, organization, and function of tight junctional complexes in the tracheal epithelium of infant ferrets.

The surface epithelium of newborn ferret airways matures rapidly in the first month of life. Prominent developmental features include a transition from predominantly non-ciliated to ciliated cells, quantitative and qualitative changes in secretion of macromolecules, and a transition from secretory to absorptive patterns of ion transport. Freeze-fracture replicas of ferret tracheal epithelium from 0 to 28 days of age exhibited progressive developmental patterns in tight junctional structure from beaded, unclosed patterns in newborns to more closed patterns at 28 days. Strand number increased while the depth of tight junctional structures and the proportion of strands exhibiting discontinuity decreased postnatally. Total transepithelial conductance, paracellular conductance, and cell size decreased over the first month. Our data suggest that changes in physiological parameters that reflect epithelial tight junction permeability can be attributed, at least in part, to maturation of this intercellular junction during the postnatal period.

Features of developing ferret tracheal epithelium: ultrastructural observations of in vivo and in vitro differentiation of ciliated cells.

Ultrastructural features of the developing, surface epithelium of ferrets from birth to 28 days of age were characterized. Progressive ciliogenesis in vivo was observed, beginning with cells covering the membranous portion of the trachea. Emerging cilia appeared in ultrathin sections and by scanning electron microscopy at sites correlating with accumulation of integral membrane particles seen in freeze-fracture preparations. Two patterns of ciliogenesis were observed: (1) the random emergence of cilia over the apical cell surface, and (2) initial emergence of cilia at the peripheral boundary of the luminal border of individual cells. Novel, ringlike structures were observed on the surfaces of nonciliated cells at all ages studied. Active ciliogenesis as well as the appearance of ring structures also were documented in the superficial epithelium from 1- to 5-day-old animals maintained in vitro for up to 4 days.

Postnatal development of tracheal surface epithelium and submucosal glands in the ferret.

We explored the usefulness of the postnatal ferret as a model for early developmental events in the large airways, using light and scanning electron microscopy. In the first 28 postnatal days, ferret tracheal surface epithelium and glands undergo dramatic growth and development. Tracheal surface area increases 8-fold. At birth, ciliated cells are sparse (9.4 +/- 1.2% of total epithelial cells). A significant increase in ciliated cells is observed at weekly intervals and by day 28 the ciliated cell is the predominant cell type (54.2 +/- 2.8% of total epithelial cells). Secretory cells decrease from 66.4 +/- 1.0% at birth to 22.2 +/- 2.8% of total epithelial cells. Histochemical staining of the granules of the epithelial secretory cells changes from predominantly non-acidic (staining with PAS but not Alcian blue) to predominantly acidic (staining also with Alcian blue). During the same time interval, tracheal glands develop from intraepithelial cellular aggregates devoid of secretory granules at birth into complex, submucosal tubuloacinar structures composed predominantly of cells containing non-acidic secretory granules at 28 days. Therefore, infant ferrets offer an opportunity to examine the structural and functional components of the mucociliary clearance mechanism at developmental stages which occur prenatally in many laboratory animals and in humans.