Functional delineation of the luminal epithelial microenvironment in breast using cell-based screening in combinatorial microenvironments.

Microenvironment signals are potent determinants of cell fate and arbiters of tissue homeostasis, however understanding how different microenvironment factors coordinately regulate cellular phenotype has been experimentally challenging. Here we used a high-throughput microenvironment microarray comprised of 2640 unique pairwise signals to identify factors that support proliferation and maintenance of primary human mammary luminal epithelial cells. Multiple microenvironment factors that modulated luminal cell number were identified, including: HGF, NRG1, BMP2, CXCL1, TGFB1, FGF2, PDGFB, RANKL, WNT3A, SPP1, HA, VTN, and OMD. All of these factors were previously shown to modulate luminal cell numbers in painstaking mouse genetics experiments, or were shown to have a role in breast cancer, demonstrating the relevance and power of our high-dimensional approach to dissect key microenvironmental signals. RNA-sequencing of primary epithelial and stromal cell lineages identified the cell types that express these signals and the cognate receptors in vivo. Cell-based functional studies confirmed which effects from microenvironment factors were reproducible and robust to individual variation. Hepatocyte growth factor (HGF) was the factor most robust to individual variation and drove expansion of luminal cells via cKit+ progenitor cells, which expressed abundant MET receptor. Luminal cells from women who are genetically high risk for breast cancer had significantly more MET receptor and may explain the characteristic expansion of the luminal lineage in those women. In ensemble, our approach provides proof of principle that microenvironment signals that control specific cellular states can be dissected with high-dimensional cell-based approaches.

A case of herpes simplex virus and cytomegalovirus retinitis coinfection.

PURPOSE: To describe a case of a 64-year-old male presenting with cytomegalovirus (CMV) and herpes simplex virus (HSV) retinitis coinfection in the setting of Burkitt's lymphoma. METHODS: Case report including multimodal imaging and anterior chamber polymerase chain reaction results. RESULTS: This case highlights the importance of the clinical exam and maintaining high diagnostic suspicion for viral retinitis in immunocompromised patients. CONCLUSIONS: Aqueous fluid PCR can be a useful adjunctive test to distinguish and confirm a diagnosis of viral retinitis. Given the limited sample volume of aqueous biopsy, it is important to prioritize the order of PCR testing based on clinical suspicion of the causative agent.

Subretinal deposits in young patients treated with voretigene neparvovec-rzyl for RPE65-mediated retinal dystrophy.

We report a series of three young patients (ages: 22 months, 2 years, and 5 years) who developed subretinal deposits at post-operative week one following subretinal voretigene neparvovec-rzyl treatment for RPE65-mediated retinal dystrophy. In the 5-year-old, subretinal deposits were also observed in the inferior periphery of both eyes. All three patients experienced improved visual function with treatment, and both the macular and inferior subretinal deposits have improved or resolved over the follow-up period. These findings may inform the delivery parameters and safety profile of AAV-based gene therapy as the number of retinal gene therapy trials continues to grow.

Desigualdades de género y las redes de apoyo social en pacientes con tuberculosis

Las desigualdades de género y las redes de apoyo social relacionadas con la tuberculosis conllevan asimetrías de poder, distintas oportunidades para gozar de salud en unos y aumentar la vulnerabilidad a enfermar en otros. El objetivo de este trabajo fue examinar las desigualdades de género según redes de apoyo social en pacientes con tuberculosis. Se realizó una revisión sistematizada en fuentes de datos digitales como Google Académico, SciELO y PudMed. Para la búsqueda se utilizaron las palabras clave: género, desigualdades, redes de apoyo y tuberculosis. Los resultados se limitaron a artículos científicos de revisión y originales, publicados en idioma español e inglés de los últimos 21 años. Se sostiene que las redes de apoyo ante un episodio mórbido resultan condicionadas por las diferencias, pero, sobre todo, por las desigualdades de género dentro de las estructuras sociales. Se concluye que la mayoría de estudios no vinculó la tuberculosis con el género. Las redes de apoyo social cumplieron con una parte fundamental en el proceso salud-enfermedad de las personas con este padecimiento.

Gender inequalities and social support networks related to tuberculosis lead to power asymmetries, health disparities for some, and increased vulnerability to disease for others. The aim of this study was to examine gender inequalities according to social support networks in patients with tuberculosis. A systematic review was conducted in digital data sources such as Google Scholar, SciELO, and PubMed. The search terms used were: gender, inequalities, support networks, and tuberculosis. The results were limited to review and original research articles, published in Spanish and English in the last 21 years. It is argued that support networks in the face of a morbid episode are conditioned by differences, but above all, by gender inequalities within social structures. It is concluded that most studies have failed to associate tuberculosis with gender. Social support networks played a fundamental role in the health-disease process of people with this disease.

Orbital schwannoma management: a case report, literature review, and potential paradigm shift.

PURPOSE: To present a case of orbital schwannoma and assess the literature on treatment modalities. METHODS: A MEDLINE literature search for cases of orbital schwannomas was performed using the PubMed search tool using the search terms "orbital schwannoma" and "orbital neurilemmoma." Papers were included if they were peer-reviewed, published in English, discussed management, and included the search terms. Each article was rated using the scale developed by the British Centre for Evidence-Based Medicine. In addition, we present a case report of an orbital schwannoma. RESULTS: A total of 428 articles were found. 102 met the criteria for inclusion. Only two articles met Level 1 evidence and 16 were important to the clinical care process. We report a case of a biopsy-proven orbital schwannoma managed conservatively with observation over a 4-year period due to risk of cosmetic disfigurement with tumor removal. There has been no change in tumor size and no associated complications during follow up. CONCLUSIONS: There is a paucity of data on the natural history of orbital schwannomas. Based on our review of the literature, we recommend observation for asymptomatic or minimally symptomatic orbital schwannomas with minimal growth over an extended period of time. For rapidly growing tumors or large tumors affecting key structures causing visual loss, diplopia, aesthetic disfigurement, or patient discomfort, a more aggressive approach may be necessary.

Incidence of Proliferative Diabetic Retinopathy and Other Neovascular Sequelae at 5 Years Following Diagnosis of Type 2 Diabetes.

OBJECTIVE: To determine the incidence and risk factors for developing proliferative diabetic retinopathy (PDR), tractional retinal detachment (TRD), and neovascular glaucoma (NVG) at 5 years after the initial diagnosis of type 2 diabetes. RESEARCH DESIGN AND METHODS: Insured patients aged ≥18 years with newly diagnosed type 2 diabetes and 5 years of continuous enrollment were identified from a nationwide commercial claims database containing data from 2007 to 2015. The incidences of PDR, TRD, and NVG were computed at 5 years following the index diagnosis of type 2 diabetes. Associations between these outcomes and demographic, socioeconomic, and medical factors were tested with multivariable logistic regression. RESULTS: At 5 years following the initial diagnosis of type 2 diabetes, 1.74% (1,249 of 71,817) of patients had developed PDR, 0.25% of patients had developed TRD, and 0.14% of patients had developed NVG. Insulin use (odds ratio [OR] 3.59, 95% CI 3.16-4.08), maximum HbA1c >9% or >75 mmol/mol (OR 2.10, 95% CI 1.54-2.69), renal disease (OR 2.68, 95% CI 2.09-3.42), peripheral circulatory disorders (OR 1.88, 95% CI 1.25-2.83), neurological disease (OR 1.62, 95% CI 1.24-2.11), and older age (age 65-74 years) at diagnosis (OR 1.62, 95% CI 1.28-2.03) were identified as risk factors for development of PDR at 5 years. Young age (age 18-23 years) at diagnosis (OR 0.46, 95% CI 0.29-0.74), Medicare insurance (OR 0.60, 95% CI 0.70-0.76), morbid obesity (OR 0.72, 95% CI 0.59-0.87), and smoking (OR 0.84, 95% CI 0.70-1.00) were identified as protective factors. CONCLUSIONS: A subset of patients with type 2 diabetes develop PDR and other neovascular sequelae within the first 5 years following the diagnosis with type 2 diabetes. These patients may benefit from increased efforts for screening and early intervention.

Deep proteome profiling of human mammary epithelia at lineage and age resolution.

Age is the major risk factor in most carcinomas, yet little is known about how proteomes change with age in any human epithelium. We present comprehensive proteomes comprised of >9,000 total proteins and >15,000 phosphopeptides from normal primary human mammary epithelia at lineage resolution from ten women ranging in age from 19 to 68 years. Data were quality controlled and results were biologically validated with cell-based assays. Age-dependent protein signatures were identified using differential expression analyses and weighted protein co-expression network analyses. Upregulation of basal markers in luminal cells, including KRT14 and AXL, were a prominent consequence of aging. PEAK1 was identified as an age-dependent signaling kinase in luminal cells, which revealed a potential age-dependent vulnerability for targeted ablation. Correlation analyses between transcriptome and proteome revealed age-associated loss of proteostasis regulation. Age-dependent proteome changes in the breast epithelium identified heretofore unknown potential therapeutic targets for reducing breast cancer susceptibility.

Changes in Immune Cell Types with Age in Breast are Consistent with a Decline in Immune Surveillance and Increased Immunosuppression.

A majority of breast cancers (BC) are age-related and we seek to determine what cellular and molecular changes occur in breast tissue with age that make women more susceptible to cancer initiation. Immune-epithelial cell interactions are important during mammary gland development and the immune system plays an important role in BC progression. The composition of human immune cell populations is known to change in peripheral blood with age and in breast tissue during BC progression. Less is known about changes in immune populations in normal breast tissue and how their interactions with mammary epithelia change with age. We quantified densities of T cells, B cells, and macrophage subsets in pathologically normal breast tissue from 122 different women who ranged in age from 24 to 74 years old. Donor-matched peripheral blood from a subset of 20 donors was analyzed by flow cytometry. Tissue immune cell densities and localizations relative to the epithelium were quantified in situ with machine learning-based image analyses of multiplex immunohistochemistry-stained tissue sections. In situ results were corroborated with flow cytometry analyses of peri-epithelial immune cells from primary breast tissue preparations and transcriptome analyses of public data from bulk tissue reduction mammoplasties. Proportions of immune cell subsets in breast tissue and donor-matched peripheral blood were not correlated. Density (cells/mm2) of T and B lymphocytes in situ decreased with age. T cells and macrophages preferentially localized near or within epithelial bilayers, rather than the intralobular stroma. M2 macrophage density was higher than M1 macrophage density and this difference was due to higher density of M2 in the intralobular stroma. Transcriptional signature analyses suggested age-dependent decline in adaptive immune cell populations and functions and increased innate immune cell activity. T cells and macrophages are so intimately associated with the epithelia that they are embedded within the bilayer, suggesting an important role for immune-epithelial cell interactions. Age-associated decreased T cell density in peri-epithelial regions, and increased M2 macrophage density in intralobular stroma suggests the emergence of a tissue microenvironment that is simultaneously immune-senescent and immunosuppressive with age.

Breast-Specific Molecular Clocks Comprised of ELF5 Expression and Promoter Methylation Identify Individuals Susceptible to Cancer Initiation.

A robust breast cancer prevention strategy requires risk assessment biomarkers for early detection. We show that expression of ELF5, a transcription factor critical for normal mammary development, is downregulated in mammary luminal epithelia with age. DNA methylation of the ELF5 promoter is negatively correlated with expression in an age-dependent manner. Both ELF5 methylation and gene expression were used to build biological clocks to estimate chronological ages of mammary epithelia. ELF5 clock-based estimates of biological age in luminal epithelia from average-risk women were within three years of chronological age. Biological ages of breast epithelia from BRCA1 or BRCA2 mutation carriers, who were high risk for developing breast cancer, suggested they were accelerated by two decades relative to chronological age. The ELF5 DNA methylation clock had better performance at predicting biological age in luminal epithelial cells as compared with two other epigenetic clocks based on whole tissues. We propose that the changes in ELF5 expression or ELF5-proximal DNA methylation in luminal epithelia are emergent properties of at-risk breast tissue and constitute breast-specific biological clocks. PREVENTION RELEVANCE: ELF5 expression or DNA methylation level at the ELF5 promoter region can be used as breast-specific biological clocks to identify women at higher than average risk of breast cancer.

Fractal Dimension Analysis of OCTA Images of Diabetic Retinopathy Using Circular Mass-Radius Method.

BACKGROUND AND OBJECTIVE: To quantify fractal dimension (FD) by mass-radius method in optical coherence tomography angiography (OCTA) images and characterize microvascular differences in eyes with and without diabetic retinopathy (DR). PATIENTS AND METHODS: A retrospective study was conducted using OCTA images of 3 mm × 3 mm and 6 mm × 6 mm scans for superficial and deep capillary plexuses from 49 control eyes and 58 eyes with DR. RESULTS: Analysis of variance showed a significant difference between the FD of control and diabetic eyes in deep plexus scans, and the 3 mm × 3 mm superficial plexus scan (P < .05). In the 3 mm × 3 mm superficial plexus, the FD of severe nonproliferative DR (NPDR) and proliferative DR (PDR) were significantly lower compared to control. The scans of the deep plexus showed only severe NPDR was significantly reduced in the 6 mm × 6 mm scan, whereas moderate NPDR, severe NPDR, and PDR were significantly lower in the 3 mm × 3 mm scan. CONCLUSION: The study suggests the use of FD as a measure of microvascular dropout in DR. [Ophthalmic Surg Lasers Imaging Retina. 2021;52:116-122.].

A structurally optimized FXR agonist, MET409, reduced liver fat content over 12 weeks in patients with non-alcoholic steatohepatitis.

BACKGROUND & AIMS: The benefits of farnesoid X receptor (FXR) agonists in patients with non-alcoholic steatohepatitis (NASH) have been validated, although improvements in efficacy and/or tolerability remain elusive. Herein, we aimed to assess the performance of a structurally optimized FXR agonist in patients with NASH. METHODS: In this 12-week, randomized, placebo-controlled study, we evaluated MET409 - a non-bile acid agonist with a unique chemical scaffold - in patients with NASH. Patients were randomized to receive either 80 mg (n = 20) or 50 mg (n = 19) of MET409, or placebo (n = 19). RESULTS: At Week 12, MET409 lowered liver fat content (LFC), with mean relative reductions of 55% (80 mg) and 38% (50 mg) vs. 6% in placebo (p <0.001). MET409 achieved ≥30% relative LFC reduction in 93% (80 mg) and 75% (50 mg) of patients vs. 11% in placebo (p <0.001) and normalized LFC (≤5%) in 29% (80 mg) and 31% (50 mg) of patients vs. 0% in placebo (p <0.05). An increase in alanine aminotransferase (ALT) was observed with MET409, confounding Week 12 changes from baseline (-25% for 80 mg, 28% for 50 mg). Nonetheless, MET409 achieved ≥30% relative ALT reduction in 50% (80 mg) and 31% (50 mg) of patients vs. 17% in placebo. MET409 was associated with on-target high-density lipoprotein cholesterol decreases (mean changes of -23.4% for 80 mg and -20.3% for 50 mg vs. 2.6% in placebo) and low-density lipoprotein cholesterol (LDL-C) increases (mean changes of 23.7% for 80 mg and 6.8% for 50 mg vs. -1.5% in placebo). Pruritus (mild-moderate) occurred in 16% (50 mg) and 40% (80 mg) of MET409-treated patients. CONCLUSION: MET409 lowered LFC over 12 weeks in patients with NASH and delivered a differentiated pruritus and LDL-C profile at 50 mg, providing the first clinical evidence that the risk-benefit profile of FXR agonists can be enhanced through structural optimization. LAY SUMMARY: Activation of the farnesoid X receptor (FXR) is a clinically validated approach for treating non-alcoholic steatohepatitis (NASH), although side effects such as itching or increases in low-density lipoprotein cholesterol are frequently dose-limiting. MET409, an FXR agonist with a unique chemical structure, led to significant liver fat reduction and delivered a favorable side effect profile after 12 weeks of treatment in patients with NASH. These results provide the first clinical evidence that the risk-benefit profile of FXR agonists can be enhanced.

Evidence for accelerated aging in mammary epithelia of women carrying germline BRCA1 or BRCA2 mutations.

During aging in the human mammary gland, luminal epithelial cells lose lineage fidelity by expressing markers normally expressed in myoepithelial cells. We hypothesize that loss of lineage fidelity is a general manifestation of epithelia that are susceptible to cancer initiation. In the present study, we show that histologically normal breast tissue from younger women who are susceptible to breast cancer, as a result of harboring a germline mutation in BRCA1, BRCA2 or PALB2 genes, exhibits hallmarks of accelerated aging. These include proportionately increased luminal epithelial cells that acquired myoepithelial markers, decreased proportions of myoepithelial cells and a basal differentiation bias or failure of differentiation of cKit+ progenitors. High-risk luminal and myoepithelial cells are transcriptionally enriched for genes of the opposite lineage, inflammatory- and cancer-related pathways. We have identified breast-aging hallmarks that reflect a convergent biology of cancer susceptibility, regardless of the specific underlying genetic or age-dependent risk or the associated breast cancer subtype.

The Protective Effect of Estrogen Against Radiation Cataractogenesis is Dependent Upon the Type of Radiation.

Astronauts participating in prolonged space missions constitute a population of individuals who are at an increased risk for cataractogenesis due to exposure to densely ionizing charged particles. Using a rat model, we have previously shown that after irradiation of eyes with either low-linear energy transfer (LET) 60Co γ rays or high-LET 56Fe particles, the rate of progression of anterior and posterior subcapsular cataracts was significantly greater in ovariectomized females implanted with 17-ß-estradiol (E2) compared to ovariectomized or intact rats. However, our additional low-LET studies indicated that cataractogenesis may be a modifiable late effect, since we have shown that the modulation of cataractogenesis is dependent upon the timing of administration of E2. Interestingly, we found that E2 protected against cataractogenesis induced by low-LET radiation, but only if administered after the exposure; if administered prior to and after irradiation, for the entire period of observation, then E2 enhanced progression and incidence of cataracts. Since most radioprotectors tested to date are unsuccessful in protecting against the effects of high-LET radiation, we wished to determine whether the protection mediated by E2 against radiation cataractogenesis induced by low-LET radiation would also be observed after high-LET irradiation. Female 56-day-old Sprague-Dawley rats were treated with E2 at various times relative to the time of single-eye irradiation with 2 Gy of 56Fe ions. We found that administration of E2 before irradiation and throughout the lifetime of the rat enhanced cataractogenesis compared to ovariectomized animals. The enhancing effect was slightly reduced when estrogen was removed after irradiation. However, in contrast to what we observed after γ-ray irradiation, there was no inhibition of cataractogenesis if E2 was administered only after 56Fe-ion irradiation. We conclude that protection against cataractogenesis by estrogen is dependent upon the type and ionization density of radiation that the lens was exposed to. The lack of inhibition of radiation cataractogenesis in rats that receive E2 treatment after high-LET irradiation may be attributed to the qualitative differences in the types of DNA damage induced with high-LET radiation compared to low-LET radiation or how damage may be modified at the DNA or tissue level after irradiation.

Text Messaging Interventions on Cancer Screening Rates: A Systematic Review.

BACKGROUND: Despite high-quality evidence demonstrating that screening reduces mortality from breast, cervical, colorectal, and lung cancers, a substantial portion of the population remains inadequately screened. There is a critical need to identify interventions that increase the uptake and adoption of evidence-based screening guidelines for preventable cancers at the community practice level. Text messaging (short message service, SMS) has been effective in promoting behavioral change in various clinical settings, but the overall impact and reach of text messaging interventions on cancer screening are unknown. OBJECTIVE: The objective of this systematic review was to assess the effect of text messaging interventions on screening for breast, cervical, colorectal, and lung cancers. METHODS: We searched multiple databases for studies published between the years 2000 and 2017, including PubMed, EMBASE, and the Cochrane Library, to identify controlled trials that measured the effect of text messaging on screening for breast, cervical, colorectal, or lung cancers. Study quality was evaluated using the Cochrane risk of bias tool. RESULTS: Our search yielded 2238 citations, of which 31 underwent full review and 9 met inclusion criteria. Five studies examined screening for breast cancer, one for cervical cancer, and three for colorectal cancer. No studies were found for lung cancer screening. Absolute screening rates for individuals who received text message interventions were 0.6% to 15.0% higher than for controls. Unadjusted relative screening rates for text message recipients were 4% to 63% higher compared with controls. CONCLUSIONS: Text messaging interventions appear to moderately increase screening rates for breast and cervical cancer and may have a small effect on colorectal cancer screening. Benefit was observed in various countries, including resource-poor and non-English-speaking populations. Given the paucity of data, additional research is needed to better quantify the effectiveness of this promising intervention.

Identification of Multiple Proteins Coupling Transcriptional Gene Silencing to Genome Stability in Arabidopsis thaliana.

Eukaryotic genomes are regulated by epigenetic marks that act to modulate transcriptional control as well as to regulate DNA replication and repair. In Arabidopsis thaliana, mutation of the ATXR5 and ATXR6 histone methyltransferases causes reduction in histone H3 lysine 27 monomethylation, transcriptional upregulation of transposons, and a genome instability defect in which there is an accumulation of excess DNA corresponding to pericentromeric heterochromatin. We designed a forward genetic screen to identify suppressors of the atxr5/6 phenotype that uncovered loss-of-function mutations in two components of the TREX-2 complex (AtTHP1, AtSAC3B), a SUMO-interacting E3 ubiquitin ligase (AtSTUbL2) and a methyl-binding domain protein (AtMBD9). Additionally, using a reverse genetic approach, we show that a mutation in a plant homolog of the tumor suppressor gene BRCA1 enhances the atxr5/6 phenotype. Through characterization of these mutations, our results suggest models for the production atxr5 atxr6-induced extra DNA involving conflicts between the replicative and transcriptional processes in the cell, and suggest that the atxr5 atxr6 transcriptional defects may be the cause of the genome instability defects in the mutants. These findings highlight the critical intersection of transcriptional silencing and DNA replication in the maintenance of genome stability of heterochromatin.

Curricular innovations for medical students in palliative and end-of-life care: a systematic review and assessment of study quality.

BACKGROUND: Recent focus on palliative and end-of-life care has led medical schools worldwide to enhance their palliative care curricula. OBJECTIVE: The objective of the study was to describe recent curricular innovations in palliative care for medical students, evaluate the quality of studies in the field, and inform future research and curricular design. METHODS: The authors searched Medline, Scopus, and Educational Resource Information Center (ERIC) for English-language articles published between 2007 and 2013 describing a palliative care curriculum for medical students. Characteristics of the curricula were extracted, and methodological quality was assessed using the Medical Education Research Study Quality Instrument (MERSQI). RESULTS: The sample described 48 curricula in 12 countries. Faculty were usually interdisciplinary. Palliative care topics included patient assessment, communication, pain and symptom management, psychosocial and spiritual needs, bioethics and the law, role in the health care system, interdisciplinary teamwork, and self-care. Thirty-nine articles included quantitative evaluation, with a mean MERSQI score of 9.9 (on a scale of 5 to 18). The domain most likely to receive a high score was data analysis (mean 2.51 out of 3), while the domains most likely to receive low scores were validity of instrument (mean 1.05) and outcomes (mean 1.31). CONCLUSIONS: Recent innovations in palliative care education for medical students represent varied settings, learner levels, instructors, educational modalities, and palliative care topics. Future curricula should continue to incorporate interdisciplinary faculty. Studies could be improved by integrating longitudinal curricula and longer-term outcomes; collaborating across institutions; using validated measures; and assessing higher-level outcomes including skills, behaviors, and impact on patient care.

Strongyloides stercolaris en lavado broncoalveolar / Strongyloides stercoralis in bronchoalveolar lavage specimen

Las estrongiloidiosis es una infección parasitaria frecuente en zonas tropicales y subtropicales. Suele ser asintomática y limitarse al intestino. Sin embargo, pueden darse casos de infección extraintestinal diseminada y potencialmente fatales en pacientes inmunocomprometidos. Se presenta el caso de una paciente diagnosticada con estrongiloidiosis mediante una muestra de lavado broncoalveolar procesada con los métodos de cytospin y citología convencional...

Strongyloidiasis is a parasitic infection found especially in tropical and subtropical regions. It is usually an asymptomatic and limited disease of the gut. However, potentially fatal cases of disseminated hyperinfection in immunosuppressed patients can occur. We present the case of a female patient with strongyloidiasis in bronchoalveolar lavage specimen processed as cytospin preparations and conventional cytology...

Identification of Mre11 as a target for heat radiosensitization.

Thermal radiosensitization is believed to be mediated by an inhibition of double-strand break (DSB) repair, but the exact mechanism of radiosensitization remains to be elucidated. Previously, we demonstrated that proteins of the Mre11/Rad50/Nbs1 complex (MRN) translocate from the nucleus to the cytoplasm in cells have that been heated or heated and then irradiated; this finding led us to propose that heat radiosensitization was due at least in part to translocation of MRN. In the current study, we used leptomycin B to inhibit MRN translocation in heated, irradiated cells, but we found that heat radiosensitization was not altered. Thus enhanced radiosensitivity was not attributed to translocation of MRN proteins. To determine which of the MRN subunits contributed to heat radiosensitization, we compared the extent of heat radiosensitization in wild-type cells with that of cells hypomorphic for Mre11 or Nbs1 or cells in which the level of Rad50 was suppressed. We found that neither Nbs1 nor Rad50 is involved in heat radiosensitization, because a similar amount of heat radiosensitization was observed in cells deficient in those proteins compared to cells expressing normal levels. However, heat radiosensitization was not observed in A-TLD1 cells deficient in Mre11. Measurement of exonuclease activity of purified Mre11 heated at 42.5°C or 45.5°C indicated that the protein is very heat-labile. Immunoprecipitation of Mre11 from heated HeLa cells also revealed that hsp70 associates with Mre11 and that this association is maintained long after heating. Taken together, these findings implicate Mre11 as a target for heat radiosensitization and suggest that heat radiosensitization and inhibition of DSB repair may be mediated by heat-induced conformational changes in Mre11.

Age and hormonal status as determinants of cataractogenesis induced by ionizing radiation. I. Densely ionizing (high-LET) radiation.

Astronauts participating in extended lunar missions or the projected mission to Mars would likely be exposed to significant doses of high-linear energy transfer (LET) heavy energetic charged (HZE) particles. Exposure to even relatively low doses of such space radiation may result in a reduced latent period for and an increased incidence of lens opacification. However, the determinants of cataractogenesis induced by densely ionizing radiation have not been clearly elucidated. In the current study, we show that age at the time of exposure is a key determinant of cataractogenesis in rats whose eyes have been exposed to 2 Gy of (56)Fe ions. The rate of progression of cataractogenesis was significantly greater in the irradiated eyes of 1-year-old rats compared to young (56-day-old) rats. Furthermore, older ovariectomized rats that received exogenous estrogen treatment (17-ß-estradiol) commencing 1 week prior to irradiation and continuing throughout the period of observation of up to approximately 600 days after irradiation showed an increased incidence of cataracts and faster progression of opacification compared to intact rats with endogenous estrogen or ovariectomized rats. The same potentiating effect (higher incidence, reduced latent period) was observed for irradiated eyes of young rats. Modulation of estrogen status in the 1-year-old animals (e.g., removal of estrogen by ovariectomy or continuous exposure to estrogen) did not increase the latent period or reduce the incidence to that of intact 56-day-old rats. Since the rapid onset and progression of cataracts in 1-year-old compared to 56-day-old rats was independent of estrogen status, we conclude that estrogen cannot account for the age-dependent differences in cataractogenesis induced by high-LET radiation.

Effects of estrogen and gender on cataractogenesis induced by high-LET radiation.

Planning for long-duration manned lunar and interplanetary missions requires an understanding of radiation-induced cataractogenesis. Previously, it was demonstrated that low-linear energy transfer (LET) irradiation with 10 Gy of (60)Co gamma rays resulted in an increased incidence of cataracts in male rats compared to female rats. This gender difference was not due to differences in estrogen, since male rats treated with the major secreted estrogen 17-beta-estradiol (E2) showed an identical increase compared to untreated males. We now compare the incidence and rate of progression of cataracts induced by high-LET radiation in male and female Sprague-Dawley rats. Rats received a single dose of 1 Gy of 600 MeV (56)Fe ions. Lens opacification was measured at 2-4-week intervals with a slit lamp. The incidence and rate of progression of radiation-induced cataracts was significantly increased in the animals in which estrogen was available from endogenous or exogenous sources. Male rats with E2 capsules implanted had significantly higher rates of progression compared to male rats with empty capsules implanted (P = 0.025) but not compared to the intact female rats. These results contrast with data obtained after low-LET irradiation and suggest the possibility that the different types of damage caused by high- and low-LET radiation may be influenced differentially by steroid sex hormones.