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PurposeTo report OCT appearance and surgical outcomes of full-thickness macular holes (MHs) accidentally caused by laser devices.Patients and methodsThis retrospective case series included 11 eyes of 11 patients with laser-induced MHs treated by pars plana vitrectomy, internal limiting membrane (ILM) peeling, and gas or silicone oil tamponade. Evaluations included a full ophthalmic examination, macular spectral-domain optical coherence tomography (SD-OCT), and fundus photography. Main outcome measures is MH closure and final visual acuity; the secondary outcome was the changes of retinal pigment epithelium and photoreceptor layer evaluated by sequential post-operative SD-OCT images.ResultsFive patients were accidentally injured by a yttrium aluminum garnet (YAG) laser and six patients by handheld laser. MH diameters ranged from 272 to 815 μm (mean, 505.5±163.0 μm) preoperatively. Best-corrected visual acuity (BCVA) improved from a mean of 0.90 logMAR (range, counting finger-8/20) preoperatively to a mean of 0.34 logMAR (range, a counting finger-20/20) postoperatively (P=0.001, t=4.521). Seven of 11 patients (63.6%) achieved a BCVA better than 10/20. Ten patients had a subfoveal hyperreflectivity and four patients had a focal choroidal depression subfoveal preoperatively. At the last follow-up, all 11 eyes demonstrated the following: closure of the macular hole, variable degrees of disruption of external limiting membrane (ELM) and outer photoreceptor ellipsoid and interdigitation bands. In 10 eyes, the disruption was in the form of focal defects in the outer retina. After surgery, the subfoveal hyperreflectivity and focal choroidal depression remained.ConclusionAccidental laser-induced full-thickness macular holes can be successfully closed with surgery. Inadvertent retinal injury from laser devices, especially handheld laser injury has occurred with increasing frequency in recent years. However, there is a paucity of data regarding these types of injuries, mostly in the form of case reports. We hereby reported 11 eyes of 11 patients with laser-induced macular holes treated by vitrectomy. All the macular holes closed after surgery and the corresponding visual acuities significantly improved postoperatively.

Sun-exposed skin is characterized by superficial changes such as wrinkles, sagging and pigmentary changes, and also many internal changes in the structure and function of epidermis, basement membrane (BM) and dermis. These changes (so-called photoageing) are predominantly induced by the ultraviolet (UV) component of sunlight. Epidermis of UV-irradiated skin produced several enzymes such as matrix metalloproteinases (MMPs), urinary plasminogen activator (uPA)/plasmin and heparanase, which degrade dermal collagen fibres and elastic fibres in the dermis, and components of epidermal BM. The BM at the dermal-epidermal junction (DEJ) controls dermal-epidermal signalling and plays an important role in the maintenance of a healthy epidermis and dermis. BM is repetitively damaged in sun-exposed skin compared with unexposed skin, leading to epidermal and dermal deterioration and accelerated skin ageing. UV exposure also induces an increase in vascular endothelial growth factor (VEGF), an angiogenic factor, while thrombospondin-1 (TSP-1), an anti-angiogenic factor, is decreased; these changes induce angiogenesis in papillary dermis with increased migration of elastase-positive leucocytes, leading to dermal elastic fibre damage. Elastic fibres, such as oxytalan fibres in papillary dermis, are associated with not only skin resilience, but also skin surface texture, and elastic fibre formation by fibroblasts is facilitated by increased expression of fibulin-5. Thus, induction of fibulin-5 expression is a damage-repair mechanism, and fibulin-5 is an early marker of photoaged skin. UV-induced skin damage is cumulative and leads to premature ageing of skin. However, appropriate daily skincare may ameliorate photoageing by inhibiting processes causing damage and enhancing repair processes., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Oral mucositis (ulcer) is a serious and painful side effect for patients with head and neck cancer following radiation therapy. However, current clinical strategies cannot efficiently prevent the occurrence of oral mucositis. In this study, we investigated whether bone marrow-derived cells (BMDCs) prevented the occurrence and/or decreased the severity of radiation-induced oral mucositis. Fresh concentrated BMDCs from male C3H mice were transplanted intravenously into female mice after tongue irradiation. For 14 days postirradiation, the changes of body weight and the time courses of ulceration were observed. Until the ulcer reached maximum size (7 days postirradiation), macroscopic and histologic analyses of harvested tongues were performed to detect the behavior of donor BMDCs. Between 2 and 5 days postirradiation, BMDCs-transplanted mice showed more expression of stem cell markers (c-Kit, Sca-1) and EGFR and fewer apoptotic cells when compared with nontransplanted control mice (irradiation group). On day 7, there were fewer and smaller ulcers observed in the BMDCs-transplanted group. Tongues of these mice had preserved their epithelial thickness, and regenerative activities (blood vessels formation, cell proliferation) were higher than they were in the irradiation group. Fluorescently labeled BMDCs were not detected in tongue epithelium but rather in connective tissue (dermis) just below the basal cell layer. These findings suggest that exogenous BMDCs behave to reduce radiogenic oral mucositis in a paracrine manner., (© International & American Associations for Dental Research.)

Solar ultraviolet (UV) B radiation is known to induce matrix metalloproteinases (MMPs) that degrade collagen in the basement membrane. Macrophage migration inhibitory factor (MIF) is a pluripotent cytokine that plays an essential role in the pathophysiology of skin inflammation induced by UV irradiation. This study examined the effects of MIF on basement membrane damage following chronic UVB irradiation in mice. The back skin of MIF transgenic (Tg) and wild-type (WT) mice was exposed to UVB three times a week for 10 weeks. There was a decrease in intact protein levels of type IV collagen and increased basement membrane damage in the exposed skin of the MIF Tg mice compared to that observed in the WT mice. Moreover, the skin of the MIF Tg mice exhibited higher MIF, MMP-2 and MMP-9 expression and protein levels than those observed in the WT mice. We also found that chronic UVB exposure in MIF Tg mice resulted in higher levels of neutrophil infiltration in the dermis compared with that observed in the WT mice. In vitro experiments revealed that MIF induced increases in the MMPs expression, including that of MMP-9 in keratinocytes and MMP-2 in fibroblasts. Cultured neutrophils also secreted MMP-9 stimulated by MIF. Therefore, MIF-mediated basement membrane damage occurs primarily through MMPs activation and neutrophil influx in murine skin following chronic UVB irradiation.

The aim of this study was to evaluate the suitability of tissue-engineered mucosa (TEM) as a model for studying the acute effects of ionizing radiation (IR) on the oral mucosa. TEM and native non-keratinizing oral mucosa (NNOM) were exposed to a single dose of 16.5Gy and harvested at 1, 6, 24, 48, and 72h post-irradiation. DNA damage induced by IR was determined using p53 binding protein 1 (53BP1), and DNA repair was determined using Rad51. Various components of the epithelial layer, basement membrane, and underlying connective tissue were analyzed using immunohistochemistry. The expression of cytokines interleukin-1β (IL-1β) and transforming growth factor beta 1 (TGF-β1) was analyzed using an enzyme-linked immunosorbent assay. The expression of DNA damage protein 53BP1 and repair protein Rad51 were increased post-irradiation. The expression of keratin 19, vimentin, collage type IV, desmoglein 3, and integrins α6 and β4 was altered post-irradiation. Proliferation significantly decreased at 24, 48, and 72h post-irradiation in both NNOM and TEM. IR increased the secretion of IL-1β, whereas TGF-β1 secretion was not altered. All observed IR-induced alterations in TEM were also observed in NNOM. Based on the similar response of TEM and NNOM to IR we consider our TEM construct a suitable model to quantify the acute biological effects of IR., (Copyright © 2013 International Association of Oral and Maxillofacial Surgeons. Published by Elsevier Ltd. All rights reserved.)

Maintenance of both normal epithelial tissues and their malignant counterparts is supported by the host tissue stroma. The tumor stroma mainly consists of the basement membrane, fibroblasts, extracellular matrix, immune cells, and vasculature. Although most host cells in the stroma possess certain tumor-suppressing abilities, the stroma will change during malignancy and eventually promote growth, invasion, and metastasis. There is growing evidence that the stroma influences importantly the response to radiation therapy (RT). On the one hand, irradiation releases numerous inflammatory cytokines within the extracellular matrix and activates tumor specific antigens presentation, triggering an immune reaction that contributes to the antitumor effect seen after RT. On the other hand, the stroma significantly contributes to radioresistance but also increases the metastatic risk. Indeed, fibroblasts, which are major actors of the impact of stroma on tumor response, are involved in activation of autocrine and paracrine molecular signaling pathways regulating tumor cell proliferation, cell death, response to hypoxia, DNA repair systems and mesenchymal-epithelial transition. They are also actors of the peritumoral desmoplastic reaction, which decreases tumor radiosensitivity. The irradiated stroma can also contribute to tumor relapse after RT through recruitment of bone marrow-derived progenitors that contribute to local tumor relapse through neovascularization. It is therefore time to question preclinical models that would not take into account this impact of stroma. The increasing knowledge of the relationship between stroma and response to IR could help developing innovative strategies for potentially improve antitumor effect of RT., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Skin exposure to ionizing radiation affects the normal wound healing process and greatly impacts the prognosis of affected individuals. We investigated the effect of ionizing radiation on wound healing in a rat model of combined radiation and wound skin injury. Using a soft X-ray beam, a single dose of ionizing radiation (10-40 Gy) was delivered to the skin without significant exposure to internal organs. At 1 h postirradiation, two skin wounds were made on the back of each rat. Control and experimental animals were euthanized at 3, 7, 14, 21 and 30 days postirradiation. The wound areas were measured, and tissue samples were evaluated for laminin 332 and matrix metalloproteinase (MMP) 2 expression. Our results clearly demonstrate that radiation exposure significantly delayed wound healing in a dose-related manner. Evaluation of irradiated and wounded skin showed decreased deposition of laminin 332 protein in the epidermal basement membrane together with an elevated expression of all three laminin 332 genes within 3 days postirradiation. The elevated laminin 332 gene expression was paralleled by an elevated gene and protein expression of MMP2, suggesting that the reduced amount of laminin 332 in irradiated skin is due to an imbalance between laminin 332 secretion and its accelerated processing by elevated tissue metalloproteinases. Western blot analysis of cultured rat keratinocytes showed decreased laminin 332 deposition by irradiated cells, and incubation of irradiated keratinocytes with MMP inhibitor significantly increased the amount of deposited laminin 332. Furthermore, irradiated keratinocytes exhibited a longer time to close an artificial wound, and this delay was partially corrected by seeding keratinocytes on laminin 332-coated plates. These data strongly suggest that laminin 332 deposition is inhibited by ionizing radiation and, in combination with slower keratinocyte migration, can contribute to the delayed wound healing of irradiated skin.

Recently, we reported that heparanase plays important roles in barrier-disrupted skin, leading to increased interaction of growth factors between epidermis and dermis and facilitating various cutaneous changes, including epidermal hyperplasia and wrinkle formation. However, the role of heparanase in sun-exposed skin remains unknown. Here, we show that heparanase in human keratinocytes is activated by ultraviolet B (UVB) exposure and that heparan sulfate of perlecan is markedly degraded in UVB-irradiated human skin. The degradation of heparan sulfate resulted in a marked reduction of binding activity of the basement membrane for vascular endothelial growth factor, fibroblast growth factor-2 and -7 at the dermal-epidermal junction. Degradation of heparan sulfate was observed not only in acutely UVB-irradiated skin, but also in skin chronically exposed to sun. Interestingly, heparan sulfate was found to be degraded in sun-exposed skin, but not in sun-protected skin. These findings suggest that chronic UVB exposure activates heparanase, leading to degradation of heparan sulfate in the basement membrane and increased growth factor interaction between epidermis and dermis. These changes may facilitate photo-aging.

Aging of sun-exposed skin is accelerated by three major environmental factors: UV radiation, dryness, and oxidation. UV radiation exposure is the most influential factor in skin aging (so-called photoaging). To find ways to protect against damage caused by UV exposure and to delay photoaging, we studied internal changes of sun-exposed skin compared with those of sun-protected skin. We found that the basement membrane (BM) at the dermal-epidermal junction (DEJ) of sun-exposed skin becomes damaged and multilayered and partly disrupted compared with that of sun-protected skin. BM plays important roles in maintaining a healthy epidermis and dermis, and repeated damage destabilizes the skin, accelerating the aging process. Matrix metalloproteinases (MMPs) and urinary plasminogen activator are increased in UV-irradiated skin. MMPs are detected in the cornified layer in sun-exposed skin, but not in sun-protected skin. Using skin-equivalent models, we found that MMPs and plasmin cause BM damage and that the reconstruction of BM is enhanced by inhibiting these proteinases, as well as by increasing the synthesis of BM components. Enhancement of BM repair mechanisms may be a useful strategy in retarding photoaging.Journal of Investigative Dermatology Symposium Proceedings (2009) 14, 2-7; doi:10.1038/jidsymp.2009.5.

A variety of antiangiogenic strategies have proven effective in preclinical tumor models, either as single agents or in combination with radiation. Clinical gains have been relatively modest, however, and questions remain regarding optimal scheduling. The objectives of the current work were to evaluate whether the sequencing of acute treatment critically affects tumor pathophysiological and therapeutic response. Axitinib (Pfizer Global Research & Development), an inhibitor that predominantly targets vascular endothelial growth factor receptors, was administered either before or after each daily radiation fraction in two human prostate xenograft tumor models. Tumors were frozen at sequential times to monitor changes in (1) vascular spacing, (2) pericyte and basement membrane coverage, and (3) hypoxia. Although similar reductions in blood vessel counts were observed with each tumor model, tumor vasculature was not functionally normalized. Instead, tumor hypoxia increased, accompanied by a progressive dissociation of pericytes and basement membranes. Ultimately, tumor growth inhibition was found to be equivalent for each of the combination schedules. These studies illustrate a clear advantage to combining axitinib with fractionated therapy but argue against an acute radiosensitization or radioprotection of either the tumor cells or tumor vasculature. Instead, post- and preirradiation daily drug administration serve equally well in supplementing the response to radiotherapy.

Selective indocyanine green (ICG) staining of the macula has recently become popular in internal limiting membrane (ILM) peeling allowing a better distinction of the ILM from the underlying retina. Clinically, the ILM seems to become stiffer after ICG staining facilitating ILM peeling for the retinal surgeon. In the present study, we tried to verify the cause of this biomechanical effect. Retinal samples of postmortem porcine eyes were treated with ICG and light and compared to samples treated in darkness using biomechanical force and elongation measurements. After ICG staining of the retina combined with a 3-min illumination, a significant increase in ultimate force by 45% and a decrease in ultimate elongation by 24% were found indicating greater stiffness of the ICG-stained ILM. Without light exposure there was no such effect suggesting a light-dependent process. The stiffening effect of ICG and light is due to a photosensitizing effect of ICG leading to collagen cross-linking of the ILM.

Gastrointestinal form is the second stage of the Acute Radiation Syndrome (ARS) with a threshold dose of 8 Gy. It represents an absolutely lethal clinical-pathological unit, enteritis necro-hemorrhagica (duodenitis, jejunitis, ileitis, respectively) with unknown causal therapy. The purpose of our study has been to evaluate the morphological changes in a model of radiation-induced enteritis in rats and estimate the significance of changes in biodosimetry. Wistar rats were randomly divided into 21 groups, 10 animals per group. Samples of the jejunum were taken 24, 48, 72, and 96 h after the whole-body gamma-irradiation with the doses of 1, 5, 10, 15, and 20 Gy, and routinely stained with hematoxylin and eosin. Five morphometric markers--intercryptal distance, enterocytal height on the top and base of villus, length of basal lamina of 10 enterocytes and enterocytal width--in irradiated rat jejunum were examined. The results were compared with sham-irradiated control group. After lethal doses of irradiation, all morphometric parameters of jejunum significantly changed. With the exception of intercryptal distance, they might be considered as suitable biodosimetric markers under these experimental conditions. Our morphometry results in radiation-induced jejunitis are in accordance with those in other studies. We were the first who quantified morphological post-irradiation changes in animal jejunum. Some of them might be used under experimental conditions. This experimental study is a predecessor of the clinical assessment of a specific marker. Under clinical practice, the sensitive biodosimetric parameter could serve as one of the guidance for evaluation of the absorbed dose in irradiated troops as well as rescue workers. This is in accordance with tasks and Standardization Agreement of the North Atlantic Treaty Organization.

We report a case of localized linear IgA dermatosis (LID). The patient suffered from herpes zoster on the right waist and received three localized ultraviolet (UV) light treatments. One month later he presented with bullae on the same site. Direct immunofluorescence showed deposition of linear IgA and weak C3 along the basement membrane zone. Indirect immunofluorescence on the salt-split human skin demonstrated that IgA antibodies were bound to the epidermal side. To our knowledge, this is the first case of localized LID induced by UV light treatment for herpes zoster. It is also the third case of LID induced by UV light.

Background: The epidermal basement membrane (BM) plays important roles in adhesion between epidermis and dermis, and in controlling epidermal differentiation. The BM has been reported to be damaged in sun-exposed skin. Although matrix metalloproteinases (MMPs) are believed to be involved in the BM damage, there is no good in vitro model for examining BM damage by MMPs or for exploring methods to protect the BM., Objectives: To examine the involvement of MMPs in BM damage and approaches to protect the BM from such damage by using an in vitro skin-equivalent (SE) model., Method: SE was prepared by culturing human keratinocytes on contracted collagen gel including human fibroblasts. MMP-1, -2, -3 and -9, laminin 5 and type IV and VII collagens were determined by specific sandwich ELISAs, and MMP-2 and MMP-9 were analysed by gelatin zymography. Histological examination of SE was also carried out., Results: Despite production of BM components such as laminin 5 and type IV and VII collagens in SEs, BM was rarely observed at the dermal-epidermal junction. Several MMPs, such as MMP-1, -2, -3 and -9, were observed to be present in conditioned media and some of them were in active forms. Tissue inhibitor of metalloproteinase (TIMP)-2 was not detected, although TIMP-1 was present. Synthetic MMP inhibitors, CGS27023A and MMP-inhibitor I, which inhibit MMP-1, -2, -3 and -9, markedly augmented deposition of laminin 5 and type IV and VII collagens at the dermal-epidermal junction, resulting in the formation of continuous epidermal BM., Conclusions: Our results indicate that MMPs are involved in the degradation of BM in SEs, and that MMP inhibitors exert a protective effect against BM damage.

Purpose: Nasolacrimal outflow obstruction has been associated with high-dose (>150 mCi) radioactive iodine (I(131)) treatment. Commonly used for thyroid cancer treatment, I(131) is effectively transported in the targeted tissue by the Na(+)/I symporter (NIS). We hypothesized that NIS is expressed in the lacrimal sac and nasolacrimal duct and that active accumulation of I(131) is responsible for the clinical observations seen in these patients., Methods: Reverse transcriptase-polymerase chain reaction and immunohistochemical analyses were used to evaluate NIS expression in both archived and fresh human tissues, Results: Reverse transcriptase-polymerase chain reaction analysis showed that NIS mRNA is present in the lacrimal sac. Immunohistochemical analysis indicated that NIS protein is expressed in the stratified columnar epithelial cells of the lacrimal sac and nasolacrimal duct. NIS protein was undetectable in the lacrimal gland, Wolfring and Krause glands, conjunctiva, canaliculus, and nasal mucosa. NIS-expressing columnar epithelial cells were absent and fibrosis was evident in the lacrimal sacs from I(131)-treated patients undergoing dacryocystorhinostomy., Conclusions: NIS is present in the lacrimal sac and nasolacrimal duct of humans, correlating to the anatomic areas of clinical obstruction that develop in patients treated with greater than 150 mCi of I(131). This suggests that NIS may be the vector of radiation-induced injury to the lacrimal system. To our knowledge, this is the first report of any ion transporter in the nasolacrimal outflow system and raises new questions as to the role the lacrimal sac plays in the modification of tears and in lacrimal outflow pathology.

Purpose: To assess the histology of the retinal surface after staining of the inner limiting membrane (ILM) with indocyanine green (ICG) followed by illumination with halogen or xenon light sources in human donor and porcine eyes., Methods: Ten eyes of six human donors and six porcine eyes were used in the study. In human donor eyes, the postmortem time varied between 7 and 38 hours, and porcine eyes were evaluated 9 hours after death. In all eyes, the vitreous was removed, and a few drops of 0.5% ICG were poured over the trephined posterior pole and carefully washed out after a period of 1 minute, with balanced salt solution. Then the stained retina was illuminated for 3 minutes with different light sources: a halogen light source of 145-W power or a xenon light source of 50-W power. Adjacent, unstained retina of each eye served as a control to assess postmortem artifacts. In two human and two porcine eyes ICG was applied without illumination. Retinal specimens were evaluated by light and electron microscopy., Results: In human eyes, severe disorganization of the innermost retina and ILM loss were observed after ICG application with subsequent illumination with the halogen light source. After illumination with the xenon light source, there was only slight vacuolization of the innermost retina, with mostly intact Muller cells. The ILM remained in situ in relation to the retinal surface. Intact cellular architecture was found in all specimens after ICG staining without subsequent illumination and control specimens of unstained retina. In porcine eyes, no impact attributable to the light source or ICG alone was noted in this experimental setting., Conclusions: These findings suggest that adverse effects of ICG at the retinal surface may depend on the light source used during vitrectomy and correlate with the emission spectrum of the different light sources. In addition, care should be taken when comparing results obtained in human eyes and porcine eyes.

Antiangiogenic therapy for solid tumors clearly destroys tumor vasculature and reduces tumor growth. As an unexpected bonus, drugs that neutralize VEGF signaling generate a "normalization window" for tumor vasculature. This occurs via the recruitment of pericytes to the tumor vasculature, an effect associated with the transient stabilization of vessels and improved oxygen delivery to hypoxic zones. The normalization process is mediated by angiopoietin-1 and matrix metalloproteinases and creates a window of opportunity for improved sensitivity to ionizing radiation and the delivery of chemotherapeutic drugs.

Skin aging involves both chronological and photoaging processes. The effects of these processes are often overlapping and include changes in both the stratified epithelium and the fibroblast-rich dermis. Wound healing is frequently delayed with aging and can result in scarring. A skin equivalent model can be used to study the role of cells and the extracellular matrix in the process of wound healing. Current studies using this model employ a full-thickness wound placed atop a nonwounded dermis to mimic a partial-thickness wound. However, a true reproducible partial-thickness wound model has yet to be described. In this study, we investigated whether a laser-wounded skin equivalent would be a useful partial-thickness wound healing model. Three lasers were compared for the ability to generate a reproducible wound: an erbium-YAG, a high-powered excimer, and a low-powered excimer laser. Reepithelialization ability was tested using newborn and adult skin keratinocytes, adult esophageal keratinocytes, and cdk4-overexpressing newborn keratinocytes. Keratinocyte compartmentalization and basement membrane formation were assessed by immunofluorescence. The erbium-YAG and high-powered excimer laser cut reproducible wounds but left the remaining surface either discolored due to thermal damage and/or ragged; keratinocytes were unable to migrate into the wound area. The low-powered excimer laser cut reproducible wounds, leaving the cut surface intact and visibly unaltered; keratinocytes reepithelialized the wound in a collagenase-dependent manner within 3 days; and return of compartmentalization and basement membrane occurred within 14 days. The laser-wounded skin equivalent is an adjustable, reproducible partial-thickness wound model where keratinocyte biology akin to in vivo can be studied, and will be useful to study the effects of aging on wound healing.

Objective: Basement membranes-(BM) influence polarization, differentiation, migration and proliferation of cell and play an important role in maintaining structural and functional tissue integrity. While BM alterations have been reported in various lesions (e.g. inflammation, tumors) of laryngeal-tracheal tissues, reports on radiogenic BM alterations are rare. External irradiation (IRR) of advanced head and neck tumors often includes "normal tissues" (tissues without cancer) of the larynx. In these normal tissues both single-cell damage (necrosis, apoptosis, functional cell death) and interstitial damage (edema, fibrosis, vascular alterations, cellular infiltrations) resulting in tissue remodeling can occur, depending on various IRR parameters. In this study, we set out to add to our knowledge on the phenotypic characterization of the radiogenic BM expression pattern in laryngo-tracheal tissues., Materials and Methods: In 63 laryngo-tracheal specimens from Wistar rats, we investigated the laminin (LA) and collagen IV (CIV) expression profile and distribution pattern depending on the IRR dose (fractionated IRR, 2 Gy/day, up to a total dose of 20, 40, or 60 Gy), the time since IRR (6 months vs 12 months) and animal age (1 year vs 1.5 years) using immunohistochemical methods, semiquantitative assessment, and multivariate analysis., Results: In specimens irradiated with more than 20 Gy, both BM constituents predominantly showed dose-dependent increases and sometimes fluctuations in staining at slight to moderate levels. The expression differed in frequency and level among the various tissue structures. In some structures there was decreased expression. In the vocalis muscle, laryngeal and esophageal nerve endings, recurrent laryngeal nerve and laryngeal and tracheal muscles, LA was detected at levels significantly stronger than in controls. BM surrounding gland structures, nerve endings of the piriform sinus and esophageal muscles showed a marked tendency towards increased LA expression. However, the BM underlying the mucosal layer of the supra- and subglottic region revealed decreasing LA immunoreaction up to 40 Gy IRR, but a distinct increase in expression at 60 Gy. In the esophageal and tracheal muscles, tracheal perichondrium, recurrent laryngeal nerve and capillaries, CIV was detected at significantly stronger levels than in the controls. The vocal ligament exhibited positive CIV immunoreactions adjacent to interstitial and infiltrate cells and CIV-positive BM condensations, resulting in increased staining of these structures. CIV reactions of laryngeal and hypopharyngeal nerve endings tended towards increased expression. In contrast, BM staining surrounding vocal muscle cells revealed significantly decreased expression. In addition, there was a tendency towards decreased expression for supraglottic, subglottic and hypopharyngeal muscle cells. Age and time since irradiation had no significant effect on staining., Conclusion: The BM constituents laminin and collagen IV showed prominent dose-dependent increases and sometimes fluctuations in expression. This expression pattern persisted up to one year after completion of the irradiation. Thus, these findings must be related to late radiation effects. The altered BM expression may play a role, at least in part, in structural (e.g. laryngeal edema) and functional (voice disorders) changes associated with irradiation of the head and neck area.

Whole body gamma-ray irradiation of rats with caesium-137 (137Cs) at embryonic day 20 induced marked reduction of the weight of the testis. Body weight and other tissues, however, seemed to remain normal. By light microscopy, complete loss of germ cells was observed in the testis. Other components, such as Sertoli cells and interstitial cells, seemed to be normal. The testes from day 8 postpartum rats contained very few spermatogonia compared with newborn rats, indicating loss of germ cells between days 0 and 8. In the adult, 137Cs-irradiated testes showed two conspicuous features other than the loss of germ cells: empty vacuolar spaces between Sertoli cells and multilayered seminiferous tubule basal laminae (lamina densa). The junctional structures (ectoplasmic specializations) between Sertoli cells, however, seemed normal. The thickness of each layer of multilayered basal laminae was the same as that of normal rats and electron-lucent layers similar to lamina lucida were interposed between them. Of the empty vacuolar spaces between Sertoli cells, basal laminae bridge the gap. The basal laminae contained laminin, type IV collagen and heparan sulphate proteoglycan evenly distributed among layers, suggesting a normal composition. Rough estimation of the amount of basal laminae deposited in 137Cs-irradiated rats indicates that it is within a range similar to that in normal testis. These features imply that Sertoli cells are, in part, determined perinatally to produce basal laminae for germ-line cells.

A number of studies indicate that matrix metalloproteinase might be involved in photoaging, but little is known about their direct contribution to ultraviolet-induced histologic and morphologic changes in the skin in vivo. This study reports the relationship between changes of matrix metalloproteinase activities and ultraviolet B-induced skin changes in hairless mouse. The role of matrix metalloproteinase in the skin changes was studied by topical application of a specific matrix metalloproteinase inhibitor. The backs of mice were exposed to ultraviolet B three times a week for 10 wk. Histologic studies showed that the basement membrane structure was damaged, with epidermal hyperplasia, in the first 2 wk of ultraviolet B irradiation, followed by the appearance of wrinkles, which gradually extended in the latter half of the ultraviolet B irradiation period. We observed enhancement of type IV collagen degradation activity, but not collagenase or matrix metalloproteinase-3 activity, in extracts of ultraviolet B-irradiated, wrinkle-bearing skin. Gelatin zymographic analysis revealed that gelatinases, matrix metalloproteinase-9 and matrix metalloproteinase-2, were significantly increased in the extract. In situ zymographic study clarified that the activity was specifically localized in whole epidermis of ultraviolet B-irradiated, wrinkled skin in comparison with normal skin. The activity was induced around the basal layer of the epidermis by a single ultraviolet exposure of at least one minimal erythema dose. Furthermore, topical application of a specific matrix metalloproteinase inhibitor, CGS27023A, inhibited ultraviolet B-induced gelatinase activity in the epidermis, and its repeated application prevented ultraviolet B-induced damage to the basement membrane, as well as epidermal hyperplasia and dermal collagen degradation. Ultraviolet B-induced wrinkles were also prevented by administration of the inhibitor. These results, taken together, suggest that ultraviolet B-induced enhancement of gelatinase activity in the skin contributes to wrinkle formation through the destruction of basement membrane structure and dermal collagen in chronically ultraviolet B-exposed hairless mouse, and thus topical application of matrix metalloproteinase inhibitors may be an effective way to prevent ultraviolet B-induced wrinkle formation.

Purpose: In this study, we examined the changes of tissue contents of trace elements and iron after total body irradiation (TBI) and their possible impact on late toxicities., Material and Methods: 20 female Wistar rats were randomly assigned to two groups - either radiation (n = 10) or control (n = 10). Rats in the radiation group received TBI of 5 Gy in a single fraction. Rats were sacrificed and tissue samples of heart, lung and kidney were taken 8 weeks after radiation. Tissue levels of zinc, copper, magnesium, manganese and iron analysis were performed with an atomic absorption spectrophotometer and suprapure grade standard solutions. One kidney of each animal was taken for electron microscopic analysis. Blood samples were collected from all animals and the blood chemistry related to kidney function was studied., Results: The kidney levels of Fe and Cu significantly increased 8 weeks after irradiation (p < 0.05). The Cu/Zn ratio did not reach statistical significance in any tissue, however in kidney, there was a tendency to rise (p = 0.08). Myocardium and lung content of trace elements and iron did not show any significant change 8 weeks after irradiation. Electron microscopic analysis showed significant injury in glomerular endothelial cells, renal tubules and thickening of basement membrane. Blood chemistry showed a significant rise in serum creatinine (p = 0.008) and calcium (p = 0.01) in the TBI group. Serum creatinine levels were 0.73 and 0.84 mg/dl, and serum calcium levels were 10.1 and 11.3 mg/dl in control and TBI groups, respectively., Conclusion: A sublethal dose of TBI causes deposition of Cu and Fe within the kidney after TBI. Deposition of these elements may have some additional role on the toxicity caused by direct radiation on the kidney., (Copyright 2003 S. Karger AG, Basel)

Purpose: To determine the UV absorbance of the corneal layers (epithelium, Bowman layer, stroma) in the 240- to 400-nm range., Methods: Consecutive slices (100 microm) of human cadaveric corneas were cut, and the UV absorbance of each sample was determined in a scanning spectrophotometer. In some cases the epithelium was scraped off and its absorbance measured separately., Results: The investigation of the UV-B absorption of consecutive corneal slices revealed evidence that UV-B absorption is 1.8 times higher in the anterior 100 microm of the human cornea than in the posterior layers. The UV absorbance of the posterior layers was uniform, showing no further structural dependence. The epithelium and Bowman layer are both effective absorbers of UV-B radiation., Conclusions: These results suggest that the anterior corneal layers are particularly important in preventing damage by UV-B radiation.

This paper summarizes quantitative in vivo laminin immunofluorescence analysis of mammary glands and skin epithelial structures from mice exposed to 1 GeV/amu Fe ions. Digital confocal microscopic images were quantified and linked to the rough "core-penumbra" Fe track physical description. Comparison to gamma-ray sparsely ionizing radiation suggested the core of the Fe track being responsible for a biological response only seen with energetic Fe particles. Conclusions for modeling in vivo responses to radiation were then implied., (c2002 COSPAR. Published by Elsevier Science Ltd. All rights reserved.)

The cutaneous microvasculature was examined by electron microscopy in order to compare its characteristics in photodamaged preauricular skin and in sun-protected postauricular sites of 15 Japanese women aged 58-81 years. The characteristic ultrastructural features of the microvasculature in photodamaged skin compared with those in sun-protected skin included dilated vessels embedded in elastin which depressed endothelial cells, vessels surrounded by a thick amorphous material composed of multiple laminations of a basement membrane-like material, and activated endothelial cells which had increased numbers of cytoplasmic organelles and pinocytotic vesicles. A novel finding of this study in photodamaged vessels was an increased formation of new vessels (angiogenesis) via two distinct pathways. In severe elastosis, activated endothelial cells with densely packed intracytoplasmic microfilaments extended large pseudopods into the elastotic material. In contrast, isolated mesenchymal cells, which possessed immature Weibel-Palade bodies, were scattered around pre-existing vessels within the Grenz zone. In some cases, many mesenchymal cells with electron-lucent cytoplasms aggregated and interconnected by cytoplasmic processes, which was followed by the formation of vascular structures. These results suggest that there are significant ultrastructural differences in vessels between photoaged and intrinsically aged facial skin and that the photodamaged microvascular system is characterized by the co-existence of regressive changes and angiogenesis.

Large radiation doses cause postradiation vascular hyperpermeability by disrupting endothelia. The cumulative sequences of small doses (fractionated radiotherapy) standard in clinical practice cause it too, but not by endothelial disruption: the mechanisms are unknown. In this study, correlated fluorescent and ultrastructural localisation of a tracer revealed the architecture, fine structure and function of microvessels in mouse AT17 tumours, before and after 42 Gy fractionated radiation. Before irradiation, tumour vascular permeability lay in the normophysiological range defined by the gut and cerebral cortex. A double barrier regulated permeability: vesicular transport through the endothelial wall required approximately 2 h and then the basement membrane charge barrier trapped tracer for 2 h longer. Irradiation abolished the double barrier: tracer passed instantly through both endothelial wall and underlying basement membrane, forming diffusion haloes around microvessels within 2-5 min. Structurally, irradiated tumour microvessels were lined by a continuous and vital endothelium with closed interendothelial junctions; endothelial basement membranes were intact, though loosened. Irradiated endothelia exhibited extremely active membrane motility and intracellular vesicle trafficking. Radiation treatment raised vascular permeability by enhancing transendothelial transcytosis, and by altering the passive filter properties of the subendothelial basement membrane. This type of vascular hyperpermeability should be susceptible to pharmacological modulation.

We previously reported that laminin immunoreactivity in mouse mammary epithelium is altered shortly after whole-body irradiation with 0.8 Gy from 600 MeV/nucleon iron ions but is unaffected after exposure to sparsely ionizing radiation. This observation led us to propose that the effect could be due to protein damage from the high ionization density of the ion tracks. If so, we predicted that it would be evident soon after radiation exposure in basement membranes of other tissues and would depend on ion fluence. To test this hypothesis, we used immunofluorescence, confocal laser scanning microscopy, and image segmentation techniques to quantify changes in the basement membrane of mouse skin epidermis. At 1 h after exposure to 1 GeV/nucleon iron ions with doses from 0.03 to 1.6 Gy, neither the visual appearance nor the mean pixel intensity of laminin in the basement membrane of mouse dorsal skin epidermis was altered compared to sham-irradiated tissue. This result does not support the hypothesis that particle traversal directly affects laminin protein integrity. However, the mean pixel intensity of laminin immunoreactivity was significantly decreased in epidermal basement membrane at 48 and 96 h after exposure to 0.8 Gy 1 GeV/nucleon iron ions. We confirmed this effect with two additional antibodies raised against affinity-purified laminin 1 and the E3 fragment of the long-arm of laminin 1. In contrast, collagen type IV, another component of the basement membrane, was unaffected. Our studies demonstrate quantitatively that densely ionizing radiation elicits changes in skin microenvironments distinct from those induced by sparsely ionizing radiation. Such effects may might contribute to the carcinogenic potential of densely ionizing radiation by altering cellular signaling cascades mediated by cell-extracellular matrix interactions.

The extent of radiogenic salivary gland damage depends on the radiation dose, the fractionation, and the localization of the gland in the radiation field. Because extracellular matrix proteins, for example collagen IV, belong to the main components of basement membranes (BM), which are considered to posses cell- and structure-regulating functions, this study set out to describe radiogenic BM changes. The staining profile of collagen IV was studied in 110 female rat mandibular glands depending on age (1 year vs. 1 1/2 years), dosage (2 Gy/d; total dosages 20, 40, or 60 Gy), the radiation field (inside vs. outside), and on the latency period (1/2 year vs. 1 year), using immunohistochemical methods. The stainings were assessed semiquantitatively and differences were evaluated using multivariate analysis. The staining pattern of the polyclonal antibody in rat tissues did not differ from the pattern found in human salivary glands. Collagen IV was detected at variable levels in the glandular and nerve tissue and in vascular walls (negative: adventitia). Irradiated tissues were stained more strongly than non-irradiated, and differences were significant in salivary glands exposed to more than 20 Gy. Age and the latency period had no significant effect on staining. The BM constituent collagen IV showed dose-dependent increasing expression analogous to the salivary gland damage, which could contribute to disturbed cell-matrix interactions following salivary gland radiation exposure. Several tissue structures may be more sensitive to possible scattered radiation. Information on the pretreatment status is mandatory in histopathological studies on the BM of salivary glands.

The structural state and transport properties of basolateral membrane of rat small intestine enterocytes after exposure to X-ray irradiation (0.5; 1.0 and 2.0 Gy) were studied. The substantional suppression of the active Ca(2+)-transport process concomitant to versatile changes of the membrane structure involving the surface sites and intramembrane protein-lipid complexes was revealed one day after irradiation. Taking into account the early obtained data on apical membrane functional disorders these results confirm that ionizing radiation in sublethal doses induces the structure-function modification of enterocyte plasma membrane affecting the function of the small intestine epithelial cells.

Background: Extracellular matrix proteins (EMP) form the main components of basement membranes (BM) and associated structures. Basement membranes influence tissue structure, exchange of substances, and cell growth and differentiation. While BM changes have been reported in malignant tumors and in inflammatory diseases, systematic studies of BM changes following irradiation are rare. Because functional and histomorphological damage to the salivary glands is a well known sequela of radiotherapy in humans, our goal was to describe possible BM changes using an experimental model and immunohistochemistry (IH)., Methods: In 59 rat mandibular glands we investigated the distribution of EMP by IH. The animals differed in age from 3 months to 2 years and pre-treatment status (irradiation vs no irradiation). The analyses were performed at selected time points after completion of external irradiation (< 4 months [M]/> or = 4-6 M/> 6 M)., Results: The polyclonal antibodies (anti-laminin [AL], anti-fibronectin [AF], anti-collagen III [AC-III], anti-collagen IV [AC-IV]) identified components of BM (laminin, collagen IV) and BM-associated structures (fibronectin, collagen III) in glandular tissue, in vascular walls, in nerve tissue, and the interstitial connective tissue. Various EMP were detected in different patterns. AL stained nerve tissue moderately and both different gland tissue structures and vascular walls (without adventitia) slightly, while excretory ducts and blood vessel adventitia were distinctly positive for AF. Fibronectin was also present in the connective tissue stroma. Significant differences were seen between irradiated and nonirradiated glands, often with generally stronger and more extensive staining in the irradiated group. For example most tissue structures showed distinctly increasing immunostaining for AL in positive correlation with increasing latency of irradiation up to 6 months. The age was of minor importance., Conclusions: Previous irradiation has to be considered when interpreting the EMP profile of salivary gland tissue, especially in studies both on chronic degenerative diseases and tumor differentiation developing in an area that has undergone radiation therapy.

Cytoskeletal, cytocontractile and basement membrane proteins were studied using the immunofluorescence technique in the parotid gland in female rats after half-side fractionated megavoltage irradiation. The non-irradiated parallel-handled parotid glands served as controls. The qualitative expression of cytoskeletal proteins remained unchanged 10 days following irradiation compared to controls, i.e. cytokeratin was observed but not vimentin, desmin or GFAP (glial fibrillary acidic proteins). Six months after irradiation the cytokeratin expression adjacent to duct lumina was clearly stronger. Actin staining was more pronounced in the periphery of the acini. Ten days after irradiation no alterations of the basal lamina proteins, laminin and fibronectin, were detected. Six months post-irradiation laminin deposits were detected in areas where the entire acini had degenerated and had been replaced by fibrosis. An increased expression of fibronectin was also observed in the stroma at that time, reflecting an increased fibrosis. In areas where the acini remained, laminin immunofluorescence was mainly found in basal laminae of normal thickness, but the mean diameter of the acini seemed to have increased. This indicates a regeneration of acini and a restructuring of the basal lamina of the parenchyma.

In this ultrastructural study, albino hairless mice were irradiated with long-wavelength ultraviolet (UVA) (340-400 nm) thrice weekly for 32 weeks for a cumulative dose of 8000 J/cm2. Biopsies were taken from these mice, from age-matched unirradiated controls, and from mice irradiated with UVB for 20-30 weeks with a cumulative dose of approximately 6-9 J/cm2. The most striking UVA-induced changes were 1) elastic fiber hyperplasia without evidence of fiber disintegration, 2) a large increase in randomly deposited microfibrils; 3) massive duplication of vascular basement membrane; 4) extensive endothelial cell damage; and 5) collagen fibers with smaller diameters but without apparent damage. By contrast, after UVB, the hyperplastic elastic fibers frequently appeared to be degraded. Microfibrils were only moderately increased and remained in an organized array. Also, unlike with UVA, the epidermal basement membrane was duplicated whereas that of the vessels was mainly spared. Collagen fibers showed evidence of dissolution. Thus, ultrastructural features provide further evidence that skin damage induced by UVA can be dissimilar to that induced by UVB.

Damage to bowel often complicates radiotherapy for abdominal and pelvic malignancy. The symptoms of chronic irradiation enteropathy, which often include intractable diarrhoea, are generally attributed to vascular injury. We have examined specimens of bowel resected from patients who had been therapeutically irradiated to assess the extent of injury to the enteric nerve plexuses. To facilitate visualisation of nerve fibres and cells of neural or neuroendocrine origin, sections were immunostained with antibodies to neuron-specific enolase or PGP 9.5, widely used markers of nerves and neurons. Electron microscopy was performed in selected cases. In 27 out of 33 specimens the number of nerve fibres in the lamina propria was obviously increased compared to that in control material. Scattered cells with the histological, immunohistochemical and ultrastructural features of ganglion cells were noted within the lamina propria in 23 of the specimens, and in 18 cases so-called neuroendocrine cells, not normally seen in this location, were also present. These radiation-induced changes in the innervation of the bowel may contribute to the symptoms of chronic radiation enteropathy.

In order to investigate how sunlight may affect naevi and lentigines, their melanocytes and the basement membrane, three irradiation protocols were applied directly to ten naevi and five lentigines on 2 subjects. Neither volunteer had sufficient naevi and lentigines to be able to use the three irradiation protocols on each of the subjects. Skin biopsies were fixed in glutaraldehyde followed by osmium tetroxide, thin-sectioned and examined in a Hitachi H-7000 transmission electron microscope. Following 14 consecutive single exposures of 3 MED of UVB or single exposures followed by 25 J/cm2 of UVA, 350 J/cm2 UVA with either 2040 or 2280 mJ/cm2 UVB, the basement membrane maintained its continuity. Melanocytes were not observed on the dermal side of the epidermal-dermal junction. UVA irradiation stimulated reinforcement of the basement membrane zone by collagen fibers. Centrioles found in melanocytes following irradiation suggest that these melanocytes maybe undergoing mitosis. Dermal fibroblasts were found to contain comparatively large quantities of melanin pigment. The pigment contained in these fibroblasts may in fact constitute an additional barrier against UV irradiation.

An argon laser was used to irradiate the otolithic organs of guinea pigs and cynomolgus monkeys. After stapedectomy, the argon laser (1.5 W x 0.5 sec/shot) irradiated the utricle or saccule without touching the sensory organs. The stapes was replaced over the oval window after irradiation. The animals used for acute observation were killed immediately for morphologic studies; those used for long-term observation were kept alive for 2, 4, or 10 weeks. Acute observation revealed that sensory and supporting cells were elevated from the basement membrane only in the irradiated area. No rupture of the membranous labyrinth was observed. Long-term observation revealed that the otolith of the macula utriculi had disappeared in 2-week specimens. The entire macula utricili had disappeared in 10-week specimens. No morphologic changes were observed in cochlea, semicircular canals, or membranous labyrinth. The saccule showed similar changes.

It has been shown that due to the effect of helium-neon laser-633 nm a variable potential propagates along the human basal complex. The duration of the signal leading front equals approximately 1 sec, while that of the trailing one approximately 3 sec, the signal amplitude approximately 60 mV. As a result of the depolarization signal propagation the membrane complex is depolarized for a long time approximately (30-60 min). Thus it is shown that the laser ray acts as a damaging agent on the basal complex--the control system of living processes.

Quantitative changes were measured and ultrastructural alterations were detected in rabbit corneas after exposure to a far ultraviolet irradiation (UV). Low dose exposure (105 - 8 x 105 erg/cm2) resulted one day later in a thicker epithelium and loss of the intercellular permeability barrier. In corneas exposed to higher doses (5 x 106 erg/cm2), the epithelium was considerably thinner one day later, whereas 4 days later it was of normal thickness and the endothelial cells became degenerated. The highest UV dose exposure used (a total of 20 x 106 erg/cm2 given in 4 equal daily doses) resulted in (1) swelling of the stroma and disorganization of the collagen fibers; (2) disappearance of the basement membrane; (3) degeneration of the endothelial cells; and (4) considerable reduction of epithelium thickness.

Irradiation injury of the mouse tongue was studied by electron and light microscopy. A specific lesion was found to be the result of edema. Epithelial compensation was seen in the form of proliferation of desmosomes and hemidesmosomes, elongation of rete ridges, and hypertrophy of cytoplasmic projections from basal cells.

In both Wistar rats and C-57 BL mice, the thickness of the glomerular basement membrane increases progressively with age. This change, which can be measured in cohorts of manageable size, produces a measurement of biological age--if one accepts the assumption that it correlates with the viability of the organism that exhibit it. Whole-body exposure to x-rays has no observable influence on this age-related change.

Stromal constituents have been examined in mouse small intestine 3 1/2 days after irradiation with either 18-20 Gy gamma rays or 10 Gy neutrons. These doses were chosen for their equivalent effect on the number of intestinal crypts found after treatment. Despite the fact that the topography of the villi, as imaged by scanning electron microscopy, was altered by treatment, with gamma irradiated villi showing lateral or horizontal collapse while neutron irradiation produced conical villi, few changes were seen in the villous stromal compartments. There were, however, ultrastructural changes observed in the stroma of the pericryptal plate. Changes common to both radiation schedules included disorganisation of the subepithelial stroma and an increase in the number of irregular processes. Some changes after irradiation, however, were not identical in the two groups. Gamma irradiation resulted in pale, foamy cytoplasmic vesicles, the separation of smooth muscle cells and changes in the structure of the luminal aspect of arterial blood vessels while neutron irradiation produced dense cytoplasmic vesicles and electron dense bodies within the substance of peripheral nerve twigs. The fact that the variation in the topography of villi after the two types of radiation is matched by changes in the deep stroma rather than within the villi themselves indicates that the stromal pericryptal plate is of importance in the structure of the villus and the extent to which the villi have varied from the normal finger shaped structure.

Changes in the cell kinetics of the follicular epithelium of the pig have been studied after irradiation with single and fractionated doses (30 fractions/39 days) of X rays and the results compared with previously published data for the epidermis. In the follicular epithelium there was an initial degenerative phase, during which the rate of cell depletion was independent of the radiation dose and the mode of administration. Evidence for repopulation was seen between the 14th and 18th days after single doses (15 or 20 Gy) and by the 28th day after the start of irradiation with fractionated doses (52.3-80.0 Gy). However, the degree of cell depletion and the subsequent rate of repopulation were independent of dose. The regenerative phase was characterized by an increased cell proliferation as indicated by an elevation of the labelling index. Islands of cells (colonies), with an appearance similar to cells in the normal follicular epithelium, were seen 18 days after a single dose of 20 Gy and 42 days after the start of fractionated irradiation. When compared with the epidermis, the follicular epithelium exhibited considerably less evidence of damage after both single and fractionated doses of X rays. There was a lower incidence of degenerate cells and reduced levels of cell depletion in the follicular epithelium, suggesting that cells from this region play an important role in the repopulation of the epidermis after high-dose irradiation.

We investigated the short-term effects of a single high-dose radiation upon transplanted MM46 tumor cells in mice by means of immunohistochemistry and electron microscopy. The irradiation induced: 1) giant cell formation from the 3rd day, 2) arrest of tumor cell mitosis in prophase and metaphase due to the disorganization of the mitotic spindles, 3) changes in immunoreactivity of laminin and cytoskeletons, and 4) multilayering of the vascular basal lamina and perivascular fibrosis. The above findings suggest a decrease in tumor cell compliance, growth and invasiveness and the potentiation of defensive host responses against vascular invasion after irradiation. The analysis of the temporal sequences of the events indicates that the time lapse between the optimal host response, tumor growth and invasion constitutes a critical period.

In radiation of the thorax, the lung has been shown to be a major dose-limiting organ. The early and late responses of the lung to radiation has been reviewed, with primary emphasis on the following cell types: type II pneumocyte, type I pneumocyte, pulmonary endothelial cell and macrophage. The earliest observable and quantifiable cellular response to radiation is exhibited by the type II pneumocytes as a decrease in lamellar bodies and a corresponding increase in surfactant content of the alveolar lavage. By 18-63 weeks following exposure, several type II cells, restored in their lamellar body population, undergo degeneration and sloughing into alveolar spaces. Type I pneumocytes generally exhibit little change, although some investigators describe alveolar denudation due to degenerating type I cells. Macrophages decrease in numbers following irradiation, returning to normal populations by 4 weeks. These changes correspond closely to the changes in alveolar lavage phospholipid phosphorus. Descriptions of radiation-induced damage to endothelial cells are variable. However, blebbing and vacuolation appear to be late developing responses, although altered permeability may be earlier in its expression. Radiation pneumonitis and fibrosis are the two major clinical and experimental responses of the lung to radiation following exposures of greater than 12 Gy. The former appears to involve type II cells, macrophages and pulmonary endothelial cells, and for the latter macrophages, fibroblasts, type II pneumocytes and the pulmonary endothelial cells are involved. The two events are not interdependent, and may not necessarily be interrelated.

The effect of aging on epidermal Langerhans cells (LC) and on their response to a single ultraviolet (UV) exposure was studied in skin biopsy specimens of healthy adults, 4 aged 22-26 yr and 7 aged 62-86 yr. In unirradiated skin, old adults had fewer LC than young adults, 5.8 +/- 1.1 versus 10.0 +/- 0.8 (mean +/- SEM) per 3 mm wide cross-section (p = .015). Following irradiation with 3 times the minimal erythema dose, recognizable LC were absent in all but 2 subjects within 24 hr. However, LC number fell less rapidly in old adults and was almost unchanged at 4 hours (5.8 +/- 1.1 versus 5.0 +/- 1.2), while in young adults LC number decreased from 10.0 +/- 0.8 to 3.3 +/- 1.3 during the same period (p less than .05). Other changes noted in both young and old subjects following irradiation included cytoplasmic vacuolization, frequent apposition of LC to severely damaged keratinocytes, and the finding of LC in the basal layer of the epidermis rather than exclusively suprabasilarly as in control sections. These data demonstrate an age-associated loss of epidermal LC and slowing of LC response to UV irradiation. UV-induced LC changes appear qualitatively similar in young and old adults and include histological evidence of cellular damage, transient association of LC with damaged keratinocytes, and possible migration of LC from the irradiated epidermis within 24 hr.

The lungs of male LAF1/J mice were locally irradiated with graded doses of radiation ranging from 5 to 13 Gy. The animals were sacrificed at 1 hr, 1 day, 1 week, 4 weeks, and 12 weeks postirradiation (PI), fixed with Ruthenium Red/Triton X-100 for demonstration of basal laminar anionic sites, and processed for electron microscopy. Sham-irradiated (0 Gy, 0 time) and untreated control groups were also processed. Sections of lungs were examined ultrastructurally and changes in both alveolar and capillary basal laminar anionic sites quantitated. A marked decrease in the number of basal laminar anionic sites was noted 1 hr PI in both alveolar and capillary basal laminae at all dose levels. The decline continued to 1 week for doses of 13 Gy and more gradually to 4 weeks following doses of 5 and 9 Gy, when the number of sites began to increase. By 12 weeks animals receiving 13 Gy were approaching normal levels while those receiving 5 or 9 Gy remained subnormal. The potential effects of the loss of proteoglycans with radiation on lung basal laminar permeability and cell-extracellular matrix interactions are discussed.

The effects of far ultraviolet light irradiation upon an amphibian cornea were studied to compare the effects observed both quantitatively and ultrastructurally with data obtained after UV irradiation of mammalian corneas. The ultimate goal of this series of investigations is the elucidation of the alterations and the regeneration mechanisms, which might reflect existing morphological diversities among the species, observed in vertebrate corneas following exposure to UV light. It was found that while the epithelial cells undergo oedema after low dose exposures and are gradually damaged after high doses of UV light, 2-4 days leter a new epithelium has been formed. Intercellular permeability is increased by low dose exposure as was detected by the penetration of Ruthenium Red into the intercellular clefts. Under these conditions desmosomal structure revealed a 21-laminar configuration. The basement membrane of the amphibian, unlike that of the mammal, does not dissolve away upon exposure but shows localized disruptions which are thought to accommodate the passage of leucocytes from stroma to epithelium. That a new basement membrane is subsequently formed is evident by the existence of extracellular and intracellular secretion granules. In comparison to irradiated rabbit corneas, this stroma remains remarkably at the same thickness following a high dose exposure although a noticeable disorganization of collagen arrangement is apparent. Finally, as in the case of the rabbit corneas, a secondary degeneration of endothelium was observed 4 days after a moderate dose exposure.

The authors studied PUVA induced histological alterations in a group of 7 patients compared with 6 control subjects of same age. The epidermal alterations were unprominent: a few necrotic keratinocytes and hyperpigmentation of basal layer melanocytes with a lentiginous pattern. Basement PAS positive membrane was in some cases desestructured, but this was reversible. In the papillary dermis there was homogenization and partial or total destruction of orceinophilic vertical fibers, these phenomena were also reversible. PUVA therapy induced aging of the skin which is dose related and depends also of the patients age. No phenomena of precancerous dysplasia were observed.

The effects of prenatal irradiation on the testis are well documented, but less is known about its effects on epididymal differentiation. Pregnant rats were irradiated on the 18th day of gestation. The increase in microfilaments and lipid inclusions in the epithelial cells, in favor of a direct radiation effect, is maximal at birth and disappears thereafter. Narrow cells and clear cells show a normal differentiation pattern. On the other hand, the principal cell maturation is largely altered. The synthesis capacities are decreased based on a reduction in the size of the Golgi apparatus and the smooth endoplasmic reticulum. The aspects of invaginations of the apical plasmalemma, coated vesicles and multivesicular bodies are not modified, suggesting normal absorption functions. The epithelial basement membranes become irregular and thicker than normal, enfolding the basal part of the epithelial cells. The basement membrane proteoglycans, demonstrated by the cationic marker polyethyleneimine, are irregularly distributed in contrast to the normal pattern. These modifications of the principal cells and the basement membrane are more prominent in the proximal epididymis. This suggests a differential maturation dependence of the epithelial cells on the luminal factors, normally secreted by the testis, and likely disturbed by prenatal irradiation which leads to germ cell degeneration, and then to a new balance in the seminiferous epithelium.