The role of thymic stromal lymphopoietin in cutaneous disorders.

Thymic Stromal Lymphopoietin (TSLP) is an important cytokine that invokes early immune responses. TSLP, an IL-7-like cytokine encoded by the TSLP gene, activates JAK1 and JAK2 signaling pathways, stimulating dendritic cells to induce inflammatory Th2 cells. This cytokine is associated with pruritus in various cutaneous disorders, particularly atopic dermatitis. Varying levels of the cytokine TSLP have been demonstrated in studies of different cutaneous disorders. Pharmacological treatment targeting TSLP has been explored recently, particularly in the realm of atopic dermatitis.This review explores the relation of TSLP to cutaneous diseases, highlighting its potential as a biomarker for monitoring disease progression in discoid lupus erythematosus (DLE). The pharmacological therapy involving TSLP is discussed, along with the potential role of TSLP promotion in the treatment of alopecia areata. This overview examines the background, structure, and functions of TSLP, with a focus on its association with cutaneous disorders and a special focus on the impact of the atopic march.

[Prescription errors and drug interactions in adults hospitalized in an intensive care unit in Barranquilla (Colombia).] / Errores de prescripción e interacciones farmacológicas en personas adultas hospitalizadas en una unidad de cuidados intensivos en Barranquilla (Colombia).

OBJECTIVE: Prescribing errors and drug-drug interactions constitute a relevant topic for health professionals in these hospital settings and for the strengthening of strategies to mitigate these errors. The aim of this article was to determine the prescribing errors and drug-drug interactions present in adult patients hospitalized in an intensive care unit in the city of Barranquilla (Colombia). METHODS: A quantitative study was conducted in which 158 medical records of adult patients who were hospitalized in an Intensive Care Unit (ICU) in the city of Barranquilla during 2019 were analyzed. Medication errors and drug-drug interactions were determined by means of the Medscape application. Statistical analysis was performed using the RStudio program, descriptive and inferential statistics were applied to the data. RESULTS: Sociodemographically, male sex prevailed, the most frequent pathological history was arterial hypertension, most patients were receiving between one±five drugs, the most common errors were related to omission of dosage, route and time of administration. Drug-drug interactions were reported in 64.5% (102) of the histories and, in terms of the level of severity of the interactions, moderate interactions predominated in 32.9% (52). CONCLUSIONS: It is evident that there is a high number of medication prescription errors in hospitalized adults, among which pharmacological interactions associated mainly with the number of medications received by individuals in the ICUs stand out.

OBJETIVO: Los errores de prescripción y las interacciones farmacológicas constituyen un tema relevante para los profesionales de salud que se encuentran en estos ámbitos hospitalarios y para el fortalecimiento de estrategias que permitan mitigar estos errores. El objetivo del artículo fue determinar los errores de prescripción e interacciones medicamentosas presentes en pacientes adultos hospitalizados en una unidad de cuidados intensivos en la ciudad de Barranquilla (Colombia). METODOS: Se realizó un estudio cuantitativo en el que se analizaron 158 historias clínicas de pacientes adultos que estuvieron hospitalizados en una Unidad de Cuidados Intensivos (UCI) de la ciudad de Barranquilla durante el año 2019. Se determinaron errores de medicación e interacciones medicamentosas por medio de la aplicación Medscape. El análisis estadístico se realizó mediante el programa RStudio, se aplicó estadística descriptiva e inferencial a los datos. RESULTADOS: Sociodemográficamente prevaleció el sexo masculino, el antecedente patológico con mayor frecuencia fue la hipertensión arterial, la mayoría de los pacientes estaban recibiendo entre uno±cinco medicamentos, los errores más comunes estaban relacionados con la omisión de la dosis, vía y horario de administración. Se reportaron interacciones medicamentosas en el 64,5% (102) de las historias y, en cuanto al nivel de gravedad de las interacciones, predominaron las moderadas en un 32,9% (52). CONCLUSIONES: Se evidencia que existe un alto número de errores de prescripción de medicamentos en los adultos hospitalizados, entre los que se destacan las interacciones farmacológicas asociadas principalmente con el número de medicamentos que reciben las personas en las UCI.

Errores de prescripción e interacciones farmacológicas en personas adultas hospitalizadas en una unidad de cuidados intensivos en Barranquilla (Colombia) / Prescription errors and drug interactions in adults hospitalized in an intensive care unit in Barranquilla (Colombia)

Fundamentos: Los errores de prescripción y las interacciones farmacológicas constituyen un tema relevante para los profesio-nales de salud que se encuentran en estos ámbitos hospitalarios y para el fortalecimiento de estrategias que permitan mitigar estoserrores. El objetivo del artículo fue determinar los errores de prescripción e interacciones medicamentosas presentes en pacientesadultos hospitalizados en una unidad de cuidados intensivos en la ciudad de Barranquilla (Colombia).Métodos: Se realizó un estudio cuantitativo en el que se analizaron 158 historias clínicas de pacientes adultos que estuvieronhospitalizados en una Unidad de Cuidados Intensivos (UCI) de la ciudad de Barranquilla durante el año 2019. Se determinaron erroresde medicación e interacciones medicamentosas por medio de la aplicaciónMedscape. El análisis estadístico se realizó mediante elprogramaRStudio, se aplicó estadística descriptiva e inferencial a los datos.Resultados: Sociodemográficamente prevaleció el sexo masculino, el antecedente patológico con mayor frecuencia fue lahipertensión arterial, la mayoría de los pacientes estaban recibiendo entre uno±cinco medicamentos, los errores más comunesestaban relacionados con la omisión de la dosis, vía y horario de administración. Se reportaron interacciones medicamentosas enel 64,5% (102) de las historias y, en cuanto al nivel de gravedad de las interacciones, predominaron las moderadas en un 32,9% (52).Conclusiones: Se evidencia que existe un alto número de errores de prescripción de medicamentos en los adultos hospitali-zados, entre los que se destacan las interacciones farmacológicas asociadas principalmente con el número de medicamentos quereciben las personas en las UCI.(AU)

Backgroud: Prescribing errors and drug-drug interactions constitute a relevant topic for health professionals in these hospitalsettings and for the strengthening of strategies to mitigate these errors. The aim of this article was to determine the prescribingerrors and drug-drug interactions present in adult patients hospitalized in an intensive care unit in the city of Barranquilla (Colombia).Methods: A quantitative study was conducted in which 158 medical records of adult patients who were hospitalized in an IntensiveCare Unit (ICU) in the city of Barranquilla during 2019 were analyzed. Medication errors and drug-drug interactions were determined bymeans of the Medscape application. Statistical analysis was performed using the RStudio program, descriptive and inferential statisticswere applied to the data.Results: Sociodemographically, male sex prevailed, the most frequent pathological history was arterial hypertension, mostpatients were receiving between one±five drugs, the most common errors were related to omission of dosage, route and time ofadministration. Drug-drug interactions were reported in 64.5% (102) of the histories and, in terms of the level of severity of theinteractions, moderate interactions predominated in 32.9% (52).Conclusions: It is evident that there is a high number of medication prescription errors in hospitalized adults, among whichpharmacological interactions associated mainly with the number of medications received by individuals in the ICUs stand out.(AU)

Composición química y actividad AChE-BuChE del aceite esencial de palo santo Bursera graveolens (Kunth) Triana & Planch de Jipijapa, Ecuador / Chemical composition and AChE-BuChE activities of the essential oil of palo santo Bursera graveolens (Kunth) Tr iana & Planch from Jipijapa, Ecuador

Abstract: This article describes the chemical composition, physical properties and acetylcholinesterase (A ChE) and butyrylcholinesterase (BuChE) activity of stem - distilled essential oil (E O ) from Bursera graveolens wood chips, Burseraceae. The plant material was acquired in Quimis (Bosque de Sancán), city of Jipijapa in the province of Manabí, coastal region o f Ecuador. Thirty - six components were identified by CG - MS, which represented 98.54% of the volatile oil. The main components were limonene (68.52%) and mentofuran (20.37%). The hydrocarbon monoterpenes constituted the most abundant fractions. The average y ield of the E O was 1.26%. Regarding the physical properties of E O , the following values were obtained: relative density (1,029 g/mL), refractive index (1,477) and specific rotation (+4,567). The E O presented IC 50 inhibition values of 47.2 and 51.9 µg/mL fo r the enzymes AChE and BuChE, respectively.

Resumen: Este artículo describe la composición química, propiedades físicas y actividad acetilcolinesterasa (AChE) y butirilcolinesterasa (BuChE) del aceite esencial (AE) destilado a vapor de astillas de madera de Bursera graveolens , Burseraceae. La materia vegetal fue adquirida en Quimis (Bosque de Sancán), ciudad de Jipijapa en la provincia de Manabí, región costera d e Ecuador. Treinta y seis componentes fueron identificados por CG - MS, que representaron al 98.54 % del aceite volátil. Los componentes principales fueron limoneno (68.52%) y mentofurano (20.37%). Los monoterpenos hidrocarburos constituyeron las fracciones m ás abundantes. El rendimiento medio del AE fue de 1.26%. Con respecto a las propiedades físicas del AE se obtuvo los siguientes valores, densidad relativa (1.029 g/mL), índice de refracción (1.477) y rotación específica (+4.567). El AE presentó valores de inhibición IC 50 de 47.2 y 51.9 µg/mL para las enzimas AChE y BuChE, respectivamente.

Nursing Telemedicine Educational Encounters: Improved Patient Satisfaction in Radiation Therapy Clinics.

BACKGROUND: Nurse-led education can improve patient satisfaction, and telemedicine has increased patient access during the COVID-19 pandemic. OBJECTIVES: The aim of this article was to investigate how nursing telemedicine educational visits influence patient satisfaction. METHODS: Patients receiving standard of care in-person education for breast cancer radiation therapy (RT) between January 2019 and June 2019 comprised the preintervention cohort. After July 2019, patients received the same information virtually and represented the postintervention cohort. Press Ganey surveys were used to evaluate patient satisfaction, t tests were performed to differentiate satisfaction scores, and f tests were calculated to determine differences in the variances of response. FINDINGS: Patient satisfaction increased in the postintervention cohort for what to expect during RT, how to manage side effects, and nurses' attentiveness to patient questions and worries. There was decreased variance in patient satisfaction in the postintervention group for quality of care received from nurses and caring manner of nurses.

Post-Hurricane Distress Scale (PHDS): Determination of General and Disorder-Specific Cutoff Scores.

The Post-Hurricane Distress Scale (PHDS) was developed to assess mental health risk in the aftermath of hurricanes. We derive both disorder-specific cutoff values and a single nonspecific cutoff for the PHDS for field use by disaster relief and mental health workers. Data from 672 adult residents of Puerto Rico, sampled 3 to 12 months after Hurricane Maria, were collected. Participants completed a five-tool questionnaire packet: PHDS, Kessler K6, Patient Health Questionnaire 9, Generalized Anxiety Disorder 7, and Post-Traumatic Stress Disorder Checklist for DSM V (PCL-5). ROC curves, AUC values, sensitivities, specificities, Youden's index, and LR+ ratios are reported. The recommended single cutoff value for the PHDS is 41, whereby a respondent with a PHDS score of 41 or above is deemed high-risk for a mental health disorder. The single field use PHDS cutoff demonstrated high specificity (0.80), an LR + ratio (2.84), and a sensitivity of 0.56. The mean ROC values of PHDS for Kessler K6, Patient Health Questionnaire 9, Generalized Anxiety Disorder 7, and PCL-5 were all above 0.74. The derived cutoff for the PHDS allows efficient assessment of respondents' and/or a community's risk status for mental health disorders in the aftermath of hurricanes and natural disasters.

Understanding the histogenesis of a HRAS-PIK3R1 co-driven metastatic metaplastic breast carcinoma associated with squamous metaplasia of lactiferous ducts.

Metaplastic breast carcinoma (MBC) represents a heterogeneous group of aggressive primary breast cancers that can show differentiation into carcinomatous and sarcomatous elements. Due to its rapid growth, this malignancy can replace precursor lesions, which remain unknown in most cases. Herein, we describe a MBC presenting as a deceptive post-biopsy hematoma. Histopathological and immunohistochemical evaluation of the primary tumor revealed a squamous cell carcinoma arising in a background of squamous metaplasia of lactiferous ducts (SMOLD). In the absence of ductal carcinoma in situ, we consider SMOLD as a nonobligatory precursor of MBC. The tumor showed 'dedifferentiation' into spindle, mucin-producing, osteoclast-like giant cell and fibromatosis-like carcinoma. Next-generation sequencing revealed the driver mutations HRASQ61R and PIK3R1c.1738_1745+2del in addition to MYH11S638L and amplification of ERCC5 and FGF14, which were potential contributors to tumor phenotype. Tumor dedifferentiation was probably facilitated by epithelial-to-mesenchymal transition (EMT) with aberrant expression of platelet and endothelial adhesion molecule-1, leading to early metastasis via hematogenous route rather than lymphatic. The co-occurrence of phosphoinositide 3-kinase and mitogen-activated protein kinase pathway abnormalities along with EMT could mediate divergent growth in breast cancer.

Anti-inflammatory drug-eluting implant model system to prevent wear particle-induced periprosthetic osteolysis.

BACKGROUND: Aseptic loosening, as a consequence of an extended inflammatory reaction induced by wear particles, has been classified as one of the most common complications of total joint replacement (TJR). Despite its high incidence, no therapeutical approach has yet been found to prevent aseptic loosening, leaving revision as only effective treatment. The local delivery of anti-inflammatory drugs to modulate wear-induced inflammation has been regarded as a potential therapeutical approach to prevent aseptic-loosening. METHODS: In this context, we developed and characterized anti-inflammatory drug-eluting TiO2 surfaces, using nanoparticles as a model for larger surfaces. The eluting surfaces were obtained by conjugating dexamethasone to carboxyl-functionalized TiO2 particles, obtained by using either silane agents with amino or mercapto moieties. RESULTS: Zeta potential measurements, thermogravimetric analysis (TGA) and drug release results suggest that dexamethasone was successfully loaded onto the TiO2 particles. Release was pH dependent and greater amounts of drug were observed from amino route functionalized surfaces. The model-system was then tested for its cytotoxic and anti-inflammatory properties in LPS-stimulated macrophages. Dexamethasone released from amino route functionalized surfaces TiO2 particles was able to decrease LPS-induced nitric oxide (NO) and TNF-a production similarly to pure DEX at the same concentration; DEX released from mercapto route functionalized surfaces was at a too low concentration to be effective. CONCLUSION: Dexamethasone released from amino functionalized titanium can offer the possibility of preventing asepting loosening of joint replacement devices.

Single-Crystal X-ray Diffraction Studies of Solvated Crystals of C60 Reveal the Intermolecular Interactions between the Component Molecules.

The solid-state structures of seven solvates of C60 (C60·4tetrachloroethylene, C60·2tetrachloroethylene, C60·3benzene, C60· n-pentane, C60·diethyl ether, C60·chlorobenzene, and C60·benzene·dichloromethane) were determined by single-crystal X-ray diffraction at low temperature. At 90 K, the fullerene and solvate components are generally well-ordered and do not show the orientational disorder that plagues similar structures determined at room temperature. Interactions between the solvate molecules and the fullerene and between adjacent C60 molecules were examined and analyzed. Van der Waals and weak charge-transfer interactions are important to help to organize the individual components in these structures. The weak Lewis acid behavior of C60, such as when it cocrystallizes with diethyl ether or chlorinated solvents, is apparent. In addition, π-stacking interactions are prevalent. The solvates of C60 reported here were frequently obtained from attempts to cocrystallize C60 with another chemical compound. Although the desired cocrystals were never formed, the unincorporated molecules influenced solvate formation.

Variação do ângulo vertical na radiografia periapical para o diagnóstico, planejamento e tratamento endodôntico / Vertical angle variation of periapical radiograph for the diagnosis, planning and endodontic treatment

Objetivo: o objetivo do presente trabalho é discutir a importância da variação do ângulo vertical na radiografía periapical, em algumas situações clínicas especiais. Métodos: foram consultadas algumas fontes com o intuito de analisar a utilidade dessa técnica na Endodontia. Resultados: a variação do ângulo vertical na radiografía periapical pode ser empregada em diversas situações clínicas na Endodontia, tais como: a sobreposição do arco zigomático sobre os ápices dos molares superiores, para visualizar ápices muito finos, em casos de fraturas radiculares transversais, para evitar sobreposições das asas dos grampos do isolamento absoluto sobre os ápices dos dentes, durante o tratamento endodôntico. Também, poder ser útil para a detecção de obstruções cervicais do canal radicular e pinos intrarradiculares curtos, em dentes portadores de coroas metálicas, onde é difícil sua visualização. Conclusões: o uso adequado da variação na angulação vertical pode auxiliar na resolução satisfatória desses casos.

Complete mitochondrial genome of Ictalurus pricei (Teleostei: Ictaluridae) and evidence of a cryptic Ictalurus species in Northwest Mexico.

The nominal Yaqui catfish, Ictalurus pricei, is a species of Ictaluridae (Siluriformes) often recorded from Northwest Mexico. Southern distribution members of the I. pricei complex in Northwest Mexico include at least one undescribed species that differs from Yaqui catfish in morphological features, herein called "Sinaloa Catfish". Sequencing of four geographical mitogenome haplotypes of Yaqui catfish and Sinaloa catfish showed geographical haplotypes of I. pricei within a clade of specific identity, close to Sinaloa catfish haplotypes. Our molecular phylogeny represents a working hypothesis supporting information on the evolutionary relationships of the Ictalurus species from Western Mexico and Western USA.

Lack of training and Comfort level with Provision of Palliative Care in Puerto Rican Emergency Departments.

OBJECTIVE: Although many institutions in the United States have incorporated palliative care practices in their emergency departments, very little has occurred in Puerto Rico. Information regarding palliative care training of emergency medicine physicians in Puerto Rico is unclear and most physicians have poor or no access to palliative care services for their patients. This study explores the perceptions and barriers encountered by practicing emergency physicians in providing palliative care in Puerto Rican Emergency Departments. METHODS: A survey was administered to physicians attending the American College of Emergency Physicians Puerto Rico Chapter annual Convention Attending physicians and residents from the University of Puerto Rico School of Medicine validated the survey tool via a "content validity" approach. Participants were asked to respond to Likert scaled statements with options that ranged from "Strongly Agree" to "Strongly Disagree". The statements addressed physician comfort level with provision of palliative care and discussion of end of life issues, as well as barriers encountered by providers such as time constraints, fear of lawsuits, and lack of access to specialists among others. RESULTS: Of the 85 physicians at the convention 59 provided surveys available for review for a response rate of 70%. Of those surveyed, 35% reported feeling some level of discomfort at providing palliative care in the ED and 39.6% agreed or strongly agreed that their lack of training in palliative care affects their ability to provide this service. In addition, 81% lack access to palliative care specialists/teams in the emergency department. However, 82.8% agreed or strongly agreed that palliative care is an important competence for emergency physicians. CONCLUSIONS: Despite recognizing palliative care as an important competence, emergency physicians in Puerto Rico reported insufficiencies in training, decreased level of comfort, and lack of access to specialists in palliative care. Efforts to enhance physician training and provide palliative care resources must be pursued in order to improve the quality of care given to patients visiting Puerto Rican Emergency departments.

Hollow chitosan/alginate nanocapsules for bioactive compound delivery.

This work aimed at the development of biodegradable nanocapsules as carriers of two bioactive compounds, 5-aminosalycilic acid and glycomacropeptide. Nanocapsules were produced through layer-by-layer (LbL) deposition of chitosan (CH) and alginate (ALG) layers on polystyrene nanoparticles. The bioactive compounds were incorporated on the third layer of the nanocapsules being its encapsulation efficiency and release behaviour evaluated. The LbL deposition process, stability, morphology and size of the multilayer nanocapsules were monitored by means of zeta potential and transmission electron microscopy (TEM). The bioactive compounds release from the CH/ALG nanocapsules was successfully described by a mathematical model (linear superimposition model - LSM), which allowed concluding that bioactive compounds release is due to both Brownian motion and the polymer relaxation of the CH/ALG layers. Final results demonstrated that the synthesized LbL hollow nanocapsules presented spherical morphology and a good capacity to encapsulate different bioactive compounds, being the best results obtained for the system containing 5-aminosalycilic acid (with an encapsulation efficiency of approximately 70%). CH/ALG multilayer nanocapsules could be a promising carrier of bioactive compounds for applications in food and pharmaceutical industries.

Formation of a Stable Complex, RuCl2(S2CPPh3)(PPh3)2, Containing an Unstable Zwitterion from the Reaction of RuCl2(PPh3)3 with Carbon Disulfide.

New insight into the complexity of the reaction of the prominent catalyst RuCl2(PPh3)3 with carbon disulfide has been obtained from a combination of X-ray diffraction and (31)P NMR studies. The red-violet compound originally formulated as a cationic π-CS2 complex, [RuCl(π-CS2)(PPh3)3]Cl, has been identified as a neutral molecule, RuCl2(S2CPPh3)(PPh3)2, which contains the unstable zwitterion S2CPPh3. In the absence of RuCl2(PPh3)3, there is no sign of a reaction between triphenylphosphine and carbon disulfide, although more basic trialkylphosphines form red adducts, S2CPR3. Despite the presence of an unstable ligand, RuCl2(S2CPPh3)(PPh3)2 is remarkably stable. It survives melting at 173-174 °C intact, is stable to air, and undergoes reversible electrochemical oxidation to form a monocation. When the reaction of RuCl2(PPh3)3 with carbon disulfide is conducted in the presence of methanol, crystals of orange [RuCl(S2CPPh3)(CS)(PPh3)2]Cl·2MeOH and yellow RuCl2(CS)(MeOH)(PPh3)2 also form. (31)P NMR studies indicate that the unsymmetrical dinuclear complex (SC)(Ph3P)2Ru(µ-Cl)3Ru(PPh3)2Cl is the initial product of the reaction of RuCl2(PPh3)3 with carbon disulfide. A path connecting the isolated products is presented.

Reversible phenol oxidation and reduction in the structurally well-defined 2-Mercaptophenol-α3C protein.

2-Mercaptophenol-α3C serves as a biomimetic model for enzymes that use tyrosine residues in redox catalysis and multistep electron transfer. This model protein was tailored for electrochemical studies of phenol oxidation and reduction with specific emphasis on the redox-driven protonic reactions occurring at the phenol oxygen. This protein contains a covalently modified 2-mercaptophenol-cysteine residue. The radical site and the phenol compound were specifically chosen to bury the phenol OH group inside the protein. A solution nuclear magnetic resonance structural analysis (i) demonstrates that the synthetic 2-mercaptophenol-α3C model protein behaves structurally as a natural protein, (ii) confirms the design of the radical site, (iii) reveals that the ligated phenol forms an interhelical hydrogen bond to glutamate 13 (phenol oxygen-carboxyl oxygen distance of 3.2 ± 0.5 Å), and (iv) suggests a proton-transfer pathway from the buried phenol OH (average solvent accessible surface area of 3 ± 5%) via glutamate 13 (average solvent accessible surface area of the carboxyl oxygens of 37 ± 18%) to the bulk solvent. A square-wave voltammetry analysis of 2-mercaptophenol-α3C further demonstrates that (v) the phenol oxidation-reduction cycle is reversible, (vi) formal phenol reduction potentials can be obtained, and (vii) the phenol-O(•) state is long-lived with an estimated lifetime of ≥180 millisecond. These properties make 2-mercaptophenol-α3C a unique system for characterizing phenol-based proton-coupled electron transfer in a low-dielectric and structured protein environment.

Reversible voltammograms and a Pourbaix diagram for a protein tyrosine radical.

Reversible voltammograms and a voltammetry half-wave potential versus solution pH diagram are described for a protein tyrosine radical. This work required a de novo designed tyrosine-radical protein displaying a unique combination of structural and electrochemical properties. The α(3)Y protein is structurally stable across a broad pH range. The redox-active tyrosine Y32 resides in a desolvated and well-structured environment. Y32 gives rise to reversible square-wave and differential pulse voltammograms at alkaline pH. The formal potential of the Y32-O(•)/Y32-OH redox couple is determined to 918 ± 2 mV versus the normal hydrogen electrode at pH 8.40 ± 0.01. The observation that Y32 gives rise to fully reversible voltammograms translates into an estimated lifetime of ≥30 ms for the Y32-O(•) state. This illustrates the range of tyrosine-radical stabilization that a structured protein can offer. Y32 gives rise to quasireversible square-wave and differential pulse voltammograms at acidic pH. These voltammograms represent the Y32 species at the upper edge of the quasirevesible range. The square-wave net potential closely approximates the formal potential of the Y32-O(•)/Y32-OH redox couple to 1,070 ± 1 mV versus the normal hydrogen electrode at pH 5.52 ± 0.01. The differential pulse voltammetry half-wave potential of the Y32-O(•)/Y32-OH redox pair is measured between pH 4.7 and 9.0. These results are described and analyzed.

Electrochemical and structural properties of a protein system designed to generate tyrosine Pourbaix diagrams.

This report describes a model protein specifically tailored to electrochemically study the reduction potential of protein tyrosine radicals as a function of pH. The model system is based on the 67-residue α(3)Y three-helix bundle. α(3)Y contains a single buried tyrosine at position 32 and displays structural properties inherent to a protein. The present report presents differential pulse voltammograms obtained from α(3)Y at both acidic (pH 5.6) and alkaline (pH 8.3) conditions. The observed Faradaic response is uniquely associated with Y32, as shown by site-directed mutagenesis. This is the first time voltammetry is successfully applied to detect a redox-active tyrosine residing in a structured protein environment. Tyrosine is a proton-coupled electron-transfer cofactor making voltammetry-based pH titrations a central experimental approach. A second set of experiments was performed to demonstrate that pH-dependent studies can be conducted on the redox-active tyrosine without introducing large-scale structural changes in the protein scaffold. α(3)Y was re-engineered with the specific aim to place the imidazole group of a histidine close to the Y32 phenol ring. α(3)Y-K29H and α(3)Y-K36H each contain a histidine residue whose protonation perturbs the fluorescence of Y32. We show that these variants are stable and well-folded proteins whose helical content, tertiary structure, solution aggregation state, and solvent-sequestered position of Y32 remain pH insensitive across a range of at least 3-4 pH units. These results confirm that the local environment of Y32 can be altered and the resulting radical site studied by voltammetry over a broad pH range without interference from long-range structural effects.

MDC1 maintains genomic stability by participating in the amplification of ATM-dependent DNA damage signals.

MDC1 functions in checkpoint activation and DNA repair following DNA damage. To address the physiological role of MDC1, we disrupted the MDC1 gene in mice. MDC1-/- mice recapitulated many phenotypes of H2AX-/- mice, including growth retardation, male infertility, immune defects, chromosome instability, DNA repair defects, and radiation sensitivity. At the molecular level, H2AX, MDC1, and ATM form a positive feedback loop, with MDC1 directly mediating the interaction between H2AX and ATM. MDC1 binds phosphorylated H2AX through its BRCT domain and ATM through its FHA domain. Through these interactions, MDC1 accumulates activated ATM flanking the sites of DNA damage, facilitating further ATM-dependent phosphorylation of H2AX and the amplification of DNA damage signals. In the absence of MDC1, many downstream ATM signaling events are defective. These results suggest that MDC1, as a signal amplifier of the ATM pathway, is vital in controlling proper DNA damage response and maintaining genomic stability.