Optical Microscopy Using the Faraday Effect Reveals in Situ Magnetization Dynamics of Magnetic Nanoparticles in Biological Samples.

The study of exogenous and endogenous nanoscale magnetic material in biology is important for developing biomedical nanotechnology as well as for understanding fundamental biological processes such as iron metabolism and biomineralization. Here, we exploit the magneto-optical Faraday effect to probe intracellular magnetic properties and perform magnetic imaging, revealing the location-specific magnetization dynamics of exogenous magnetic nanoparticles within cells. The opportunities enabled by this method are shown in the context of magnetic hyperthermia; an effect where local heating is generated in magnetic nanoparticles exposed to high-frequency AC magnetic fields. Magnetic hyperthermia has the potential to be used as a cellular-level thermotherapy for cancer, as well as for other biomedical applications that target heat-sensitive cellular function. However, previous experiments have suggested that the cellular environment modifies the magnetization dynamics of nanoparticles, thus dramatically altering their heating efficiency. By combining magneto-optical and fluorescence measurements, we demonstrate a form of biological microscopy that we used here to study the magnetization dynamics of nanoparticles in situ, in both histological samples and living cancer cells. Correlative magnetic and fluorescence imaging identified aggregated magnetic nanoparticles colocalized with cellular lysosomes. Nanoparticles aggregated within these lysosomes displayed reduced AC magnetic coercivity compared to the same particles measured in an aqueous suspension or aggregated in other areas of the cells. Such measurements reveal the power of this approach, enabling investigations of how cellular location, nanoparticle aggregation, and interparticle magnetic interactions affect the magnetization dynamics and consequently the heating response of nanoparticles in the biological milieu.

Gender-inclusive practice in pregnancy determination for transgender, gender diverse and non-binary patients in medical imaging.

In recent years, there has been an increased awareness and understanding of the varying gender identities within our society. Consequently, there has also been a need for healthcare providers to be cognizant of the unique needs of a gender-diverse population. Determining the pregnancy status of transgender, gender-diverse and non-binary patients in medical imaging settings has been poorly handled, and there is a lack of standardisation in the Australian and Aotearoa New Zealand setting. The potential risk of exposing a gender-diverse pregnant patient to ionising radiation increases the need for guidance to ensure potentially pregnant persons are not missed during screening questionnaires. This review article explores various approaches to pregnancy status determination for gender-diverse patients, recognising the complexities involved and emphasising the need for future work to establish a widely accepted solution.

INTRAGASTRIC BALLOON AND IMPACT ON WEIGHT LOSS: EXPERIENCE IN QUITO, EQUADOR.

BACKGROUND: Obesity is associated with different medical conditions, such as cardiologic, respiratory, gastrointestinal, and genitourinary, and constitutes a severe health problem. AIMS: This study aimed to evaluate the use of intragastric fluid-filled balloon in the reduction of weight and other measurements related to body composition. METHODS: This is a retrospective, monocentric study involving all patients who opted for the intragastric balloon Spatz® placement from January 2018 to July 2019, with fulfillment of inclusion and exclusion criteria. The patients were analyzed after 6 and 12 months after the intragastric fluid-filled balloon placed. RESULTS: A total of 121 subjects were included in this study, with 83 (68.6%) females and 38 (31.4%) males. The mean age was 36 years and height was 1.64±0.09. Weight mean and standard deviation was 89.85±14.65 kg, and body mass index was 33.05±4.03; body mass index decreased to 29.4 kg/m2 with a mean weight of 79.83 kg, after 12 months of follow-up. There were statistical differences between body mass index and the 12 months in fat percentage, fat-free mass (kg), visceral fat area, and basal metabolic rate. There was a significant variation according to gender, with males having highest reduction. The percentage of excess weight loss was 46.19, and the total weight loss was 9.24 at the end of the study. CONCLUSIONS: The study demonstrated a benefit of intragastric fluid-filled balloon on weight loss after 12 months. At the end of treatment, body mass index and the measurements of body composition were significantly lower. Men benefited more than women from the treatment.

Intragastric balloon and impact on weight loss: experience in quito, equador / Balão intragástrico e impacto na perda de peso: experiência em quito, equador

ABSTRACT BACKGROUND: Obesity is associated with different medical conditions, such as cardiologic, respiratory, gastrointestinal, and genitourinary, and constitutes a severe health problem. AIMS: This study aimed to evaluate the use of intragastric fluid-filled balloon in the reduction of weight and other measurements related to body composition. METHODS: This is a retrospective, monocentric study involving all patients who opted for the intragastric balloon Spatz® placement from January 2018 to July 2019, with fulfillment of inclusion and exclusion criteria. The patients were analyzed after 6 and 12 months after the intragastric fluid-filled balloon placed. RESULTS: A total of 121 subjects were included in this study, with 83 (68.6%) females and 38 (31.4%) males. The mean age was 36 years and height was 1.64±0.09. Weight mean and standard deviation was 89.85±14.65 kg, and body mass index was 33.05±4.03; body mass index decreased to 29.4 kg/m2 with a mean weight of 79.83 kg, after 12 months of follow-up. There were statistical differences between body mass index and the 12 months in fat percentage, fat-free mass (kg), visceral fat area, and basal metabolic rate. There was a significant variation according to gender, with males having highest reduction. The percentage of excess weight loss was 46.19, and the total weight loss was 9.24 at the end of the study. CONCLUSIONS: The study demonstrated a benefit of intragastric fluid-filled balloon on weight loss after 12 months. At the end of treatment, body mass index and the measurements of body composition were significantly lower. Men benefited more than women from the treatment.

RESUMO RACIONAL: A obesidade está associada a diferentes condições médicas, tais como cardiológicas, respiratórias, gastrointestinais, geniturinárias entre outras e constituem um grave problema de saúde. OBJETIVOS: Avaliar o emprêgo do balão intragástrico na redução de peso e em outras medidas relacionadas à composição corporal. MÉTODOS: Estudo retrospectivo, monocêntrico, incluindo todos os pacientes que optaram pela colocação de balão intragástrico Spatz® entre janeiro de 2018 e julho de 2019, com cumprimento dos critérios de inclusão e exclusão. RESULTADOS: Cento e vinte e um indivíduos foram recrutados neste estudo. A média de idade foi de 36 anos e estatura de 1,64±0,09, sendo 83 (68,6%) do sexo feminino e 38 (31,4%) do sexo masculino. A média do peso e o desvio padrão foram de 89,85±14,65 kg e o índice de massa corporal foi de 33,05±4,03. Após 12 meses, o índice de massa corporal diminuiu para 29,4 kg/m2 com um peso médio de 79,83 kg. Foram registradas diferenças estatísticas no índice de massa corporal, no percentual de gordura, massa livre de gordura (kg), área de gordura visceral e taxa metabólica basal. Houve variação significativa de acordo com o sexo, sendo o masculino com maior redução. O porcentual de perda de excesso de peso foi de 46,19% e de perda de peso total de 9,24 %ao final do estudo. CONCLUSÕES: O estudo demonstrou benefícios do balão intragástrico na perda de peso após 12 meses de colocação do balão. Ao final do tratamento, o índice de massa corporal e as medidas de composição corporal foram significativamente menores. Os homens se beneficiaram mais do que as mulheres do tratamento.

Clot-targeted magnetic hyperthermia permeabilizes blood clots to make them more susceptible to thrombolysis.

BACKGROUND: Thrombolysis is a frontline treatment for stroke, which involves the application of tissue plasminogen activator (tPA) to trigger endogenous clot-degradation pathways. However, it is only effective within 4.5 h of symptom onset because of clot contraction preventing tPA permeation into the clot. Magnetic hyperthermia (MH) mediated by tumor-targeted magnetic nanoparticles is used to treat cancer by using local heat generation to trigger apoptosis of cancer cells. OBJECTIVES: To develop clot-targeting magnetic nanoparticles to deliver MH to the surface of human blood clots, and to assess whether this can improve the efficacy of thrombolysis of contracted blood clots. METHODS: Clot-targeting magnetic nanoparticles were developed by functionalizing iron oxide nanoparticles with an antibody recognizing activated integrin αIIbß3 (PAC-1). The magnetic properties of the PAC-1-tagged magnetic nanoparticles were characterized and optimized to deliver clot-targeted MH. RESULTS: Clot-targeted MH increases the efficacy of tPA-mediated thrombolysis in contracted human blood clots, leading to a reduction in clot weight. MH increases the permeability of the clots to tPA, facilitating their breakdown. Scanning electron microscopy reveals that this effect is elicited through enhanced fibrin breakdown and triggering the disruption of red blood cells on the surface of the clot. Importantly, endothelial cells viability in a three-dimensional blood vessel model is unaffected by exposure to MH. CONCLUSIONS: This study demonstrates that clot-targeted MH can enhance the thrombolysis of contracted human blood clots and can be safely applied to enhance the timeframe in which thrombolysis is effective.

Learning Process Effectiveness During the COVID-19 Pandemic: Teleproctoring Advanced Endoscopic Skills by Training Endoscopists in Endoscopic Sleeve Gastroplasty Procedure.

BACKGROUND AND AIMS: The COVID-19 pandemic has led health institutions to cancel many of the activities including training in different fields. Most practices and training programs have been encouraged to use teleproctoring as an alternative method to enhance physician's ability and assure training. We aimed to evaluate remote training program for endoscopy sleeve gastroplasty (ESG). METHODS: Ten consecutive patients underwent an endoscopic sleeve gastroplasty procedure guided by a proctor expert using an online platform. A stepwise approach was created to assure skill acquisition. RESULTS: All cases were safely performed with no serious adverse events under teleproctoring. The average surgical and suturing times significantly decreased during the training model. From the first 5 cases to the last 5 ones, the endoscopic procedure time decreased from 120 to 93.4 min while suturing time from 92.8 to 68.4 min. The effect size was large in both cases, and the changes were meaningful according to the fitted learning curves. CONCLUSIONS: The proposed teleproctoring program was effective to deliver advanced endoscopic skills such as endosuturing for ESG, despite the restrictions imposed by the COVID-19 pandemic.

Continuous Intraoperative Neurophysiological Monitoring of the Motor Pathways Using Depth Electrodes During Surgical Resection of an Epileptogenic Lesion: A Novel Technique.

BACKGROUND AND IMPORTANCE: Intraoperative neurophysiological monitoring of the motor pathways during epilepsy surgery is essential to safely achieve maximal resection of the epileptogenic zone. Motor evoked potential (MEP) recording is usually performed intermittently during resection using a handheld stimulator or continuously through an electrode array placed on the motor cortex. We present a novel variation of continuous MEP acquisition through previously implanted depth electrodes in the perirolandic cortex. CLINICAL PRESENTATION: A 60-yr-old woman with a history of a left frontal meningioma (World Health Organization [WHO] grade II) treated with surgical resection and radiation presented with residual right hemiparesis and refractory epilepsy. Imaging demonstrated a perirolandic lesion with surrounding edema and mass effect in the prior surgical site, suspicious for radiation necrosis versus tumor recurrence. Presurgical electrocorticography (ECoG) with orthogonal, stereotactically implanted depth electrodes (stereoelectroencephalography [SEEG]) of the perirolandic cortex captured seizure onsets from the supplementary motor area (SMA) and primary motor cortex (PMC). The patient underwent a left frontal craniotomy for repeat resection and tissue diagnosis. Intraoperative ECoG and MEPs were obtained continuously with direct cortical stimulation through the indwelling SEEG electrodes in the PMC. Maximal resection was achieved with preservation of direct cortical MEPs and without deterioration of her baseline hemiparesis. Biopsy revealed radiation necrosis. At 30-mo follow-up, the patient had only rare seizures (Engel class IIB). CONCLUSION: Intraoperative cortical MEP acquisition through implanted SEEG electrode arrays is a potentially safe and effective alternative approach to continuously monitor the motor pathways during the resection of a perirolandic epileptogenic lesion, without the need for surgical interruptions.

Involvement of Mesenchymal Stem Cells in Oral Mucosal Bacterial Immunotherapy.

Recent clinical observations indicate that bacterial vaccines induce cross-protection against infections produced by different microorganisms. MV130, a polyvalent bacterial sublingual preparation designed to prevent recurrent respiratory infectious diseases, reduces the infection rate in patients with recurrent respiratory tract infections. On the other hand, mesenchymal stem cells (MSCs) are key cell components that contribute to the maintenance of tissue homeostasis and exert both immunostimulatory and immunosuppressive functions. Herein, we study the effects of MV130 in human MSC functionality as a potential mechanism that contributes to its clinical benefits. We provide evidence that during MV130 sublingual immunization of mice, resident oral mucosa MSCs can take up MV130 components and their numbers remain unchanged after vaccination, in contrast to granulocytes that are recruited from extramucosal tissues. MSCs treated in vitro with MV130 show an increased viability without affecting their differentiation potential. In the short-term, MSC treatment with MV130 induces higher leukocyte recruitment and T cell expansion. In contrast, once T-cell activation is initiated, MV130 stimulation induces an up-regulated expression of immunosuppressor factors in MSCs. Accordingly, MV130-primed MSCs reduce T lymphocyte proliferation, induce the differentiation of dendritic cells with immunosuppressive features and favor M2-like macrophage polarization, thus counterbalancing the immune response. In addition, MSCs trained with MV130 undergo functional changes, enhancing their immunomodulatory response to a secondary stimulus. Finally, we show that MSCs are able to uptake, process and retain a reservoir of the TLR ligands derived from MV130 digestion which can be subsequently transferred to dendritic cells, an additional feature that also may be associated to trained immunity.

Esterase-Cleavable 2D Assemblies of Magnetic Iron Oxide Nanocubes: Exploiting Enzymatic Polymer Disassembling To Improve Magnetic Hyperthermia Heat Losses.

Here, we report a nanoplatform based on iron oxide nanocubes (IONCs) coated with a bioresorbable polymer that, upon exposure to lytic enzymes, can be disassembled increasing the heat performances in comparison with the initial clusters. We have developed two-dimensional (2D) clusters by exploiting benchmark IONCs as heat mediators for magnetic hyperthermia and a polyhydroxyalkanoate (PHA) copolymer, a biodegradable polymer produced by bacteria that can be digested by intracellular esterase enzymes. The comparison of magnetic heat performance of the 2D assemblies with 3D centrosymmetrical assemblies or single IONCs emphasizes the benefit of the 2D assembly. Moreover, the heat losses of 2D assemblies dispersed in water are better than the 3D assemblies but worse than for single nanocubes. On the other hand, when the 2D magnetic beads (2D-MNBs) are incubated with the esterase enzyme at a physiological temperature, their magnetic heat performances began to progressively increase. After 2 h of incubation, specific absorption rate values of the 2D assembly double the ones of individually coated nanocubes. Such an increase can be mainly correlated to the splitting of the 2D-MNBs into smaller size clusters with a chain-like configuration containing few nanocubes. Moreover, 2D-MNBs exhibited nonvariable heat performances even after intentionally inducing their aggregation. Magnetophoresis measurements indicate a comparable response of 3D and 2D clusters to external magnets (0.3 T) that is by far faster than that of single nanocubes. This feature is crucial for a physical accumulation of magnetic materials in the presence of magnetic field gradients. This system is the first example of a nanoplatform that, upon exposure to lytic enzymes, such as those present in a tumor environment, can be disassembled from the initial 2D-MNB organization to chain-like assemblies with clear improvement of the heat magnetic losses resulting in better heat dissipation performances. The potential application of 2D nanoassemblies based on the cleavable PHAs for preserving their magnetic losses inside cells will benefit hyperthermia therapies mediated by magnetic nanoparticles under alternating magnetic fields.

Fe2+ Deficiencies, FeO Subdomains, and Structural Defects Favor Magnetic Hyperthermia Performance of Iron Oxide Nanocubes into Intracellular Environment.

Herein, by studying a stepwise phase transformation of 23 nm FeO-Fe3O4 core-shell nanocubes into Fe3O4, we identify a composition at which the magnetic heating performance of the nanocubes is not affected by the medium viscosity and aggregation. Structural and magnetic characterizations reveal the transformation of the FeO-Fe3O4 nanocubes from having stoichiometric phase compositions into Fe2+-deficient Fe3O4 phases. The resultant nanocubes contain tiny compressed and randomly distributed FeO subdomains as well as structural defects. This phase transformation causes a 10-fold increase in the magnetic losses of the nanocubes, which remain exceptionally insensitive to the medium viscosity as well as aggregation unlike similarly sized single-phase magnetite nanocubes. We observe that the dominant relaxation mechanism switches from Néel in fresh core-shell nanocubes to Brownian in partially oxidized nanocubes and once again to Néel in completely treated nanocubes. The Fe2+ deficiencies and structural defects appear to reduce the magnetic energy barrier and anisotropy field, thereby driving the overall relaxation into Néel process. The magnetic losses of these nanoparticles remain unchanged through a progressive internalization/association to ovarian cancer cells. Moreover, the particles induce a significant cell death after being exposed to hyperthermia treatment. Here, we present the largest heating performance that has been reported to date for 23 nm iron oxide nanoparticles under intracellular conditions. Our findings clearly demonstrate the positive impacts of the Fe2+ deficiencies and structural defects in the Fe3O4 structure on the heating performance into intracellular environment.

Dynamical Magnetic Response of Iron Oxide Nanoparticles Inside Live Cells.

Magnetic nanoparticles exposed to alternating magnetic fields have shown a great potential acting as magnetic hyperthermia mediators for cancer treatment. However, a dramatic and unexplained reduction of the nanoparticle magnetic heating efficiency has been evidenced when nanoparticles are located inside cells or tissues. Recent studies suggest the enhancement of nanoparticle clustering and/or immobilization after interaction with cells as possible causes, although a quantitative description of the influence of biological matrices on the magnetic response of magnetic nanoparticles under AC magnetic fields is still lacking. Here, we studied the effect of cell internalization on the dynamical magnetic response of iron oxide nanoparticles (IONPs). AC magnetometry and magnetic susceptibility measurements of two magnetic core sizes (11 and 21 nm) underscored differences in the dynamical magnetic response following cell uptake with effects more pronounced for larger sizes. Two methodologies have been employed for experimentally determining the magnetic heat losses of magnetic nanoparticles inside live cells without risking their viability as well as the suitability of magnetic nanostructures for in vitro hyperthermia studies. Our experimental results-supported by theoretical calculations-reveal that the enhancement of intracellular IONP clustering mainly drives the cell internalization effects rather than intracellular IONP immobilization. Understanding the effects related to the nanoparticle transit into live cells on their magnetic response will allow the design of nanostructures containing magnetic nanoparticles whose dynamical magnetic response will remain invariable in any biological environments, allowing sustained and predictable in vivo heating efficiency.

BSA-coated magnetic nanoparticles for improved therapeutic properties.

In recent years, magnetic nanoparticles have been widely investigated due to their potential in biomedical applications. For successful in vivo applications, magnetic nanoparticles must satisfy several requirements such as biocompatibility, invisibility to the immune system, high colloidal stability in biological fluids, and long blood circulation times. In this study, we have developed a formulation in which the magnetic nanoparticles are coated with bovine serum albumin to provide enhanced colloidal stability in biological fluids preserving their magnetic properties. In addition, the nanoparticles carry a chemotherapeutic drug, showing their potential as drug delivery systems. Our results reveal the influence of protein adsorption on the colloidal stability and the dynamical magnetic response of functionalized magnetic nanoparticles. Moreover, cellular internalization and in vitro cytotoxic activity in Panc-1 pancreatic cancer cells reveal enhanced cellular internalization, successful intracellular drug delivery, and efficient anticancer activity.

Lack of renal dopamine D5 receptors promotes hypertension.

Disruption of the dopamine D(5) receptor gene in mice increases BP and causes salt sensitivity. To determine the role of renal versus extrarenal D(5) receptors in BP regulation, we performed cross-renal transplantation experiments. BP was similar between wild-type mice and wild-type mice transplanted with wild-type kidneys, indicating that the transplantation procedure did not affect BP. BP was lower among D(5)(-/-) mice transplanted with wild-type kidneys than D(5)(-/-) kidneys, demonstrating that the renal D(5) receptors are important in BP control. BP was higher in wild-type mice transplanted with D(5)(-/-) kidneys than wild-type kidneys but not significantly different from syngenic transplanted D(5)(-/-) mice, indicating the importance of the kidney in the development of hypertension. On a high-salt diet, all mice with D(5)(-/-) kidneys excreted less sodium than mice with wild-type kidneys. Transplantation of a wild-type kidney into a D(5)(-/-) mouse decreased the renal expression of AT(1) receptors and Nox-2. Conversely, transplantation of a D(5)(-/-) kidney into a wild-type mouse increased the expression of both, suggesting that both renal and extrarenal factors are important in the regulation of AT(1) receptor and Nox-2 expression. These results highlight the role of renal D(5) receptors in BP homeostasis and the pathogenesis of hypertension.

Reciprocal modulation of function between the D1 and D2 dopamine receptors and the Na+,K+-ATPase.

It is well documented that dopamine can increase or decrease the activity of the Na+,K+-ATPase (NKA, sodium pump) in an organ-specific fashion. This regulation can occur, at least partially, via receptor-mediated second messenger activation and can promote NKA insertion or removal from the plasma membrane. Using co-immunoprecipitation and mass spectrometry, we now show that, in both brain and HEK293T cells, D1 and D2 dopamine receptors (DARs) can exist in a complex with the sodium pump. To determine the impact of NKA on DAR function, biological assays were conducted with NKA and DARs co-expressed in HEK293T cells. In this system, expression of NKA dramatically decreased D1 and D2 DAR densities with a concomitant functional decrease in DAR-mediated regulation of cAMP levels. Interestingly, pharmacological inhibition of endogenous or overexpressed NKA enhanced DAR function without altering receptor number or localization. Similarly, DAR function was also augmented by small interfering RNA reduction of the endogenous NKA. These data suggest that, under basal conditions, NKA negatively regulates DAR function via protein-protein interactions. In reciprocal fashion, expression of DARs decreases endogenous NKA function in the absence of dopamine, implicating DAR proteins as regulators of NKA activity. Notably, dopamine stimulation or pertussis toxin inhibition of D2 receptor signaling did not alter NKA activity, indicating that the D2-mediated decrease in NKA function is dependent upon protein-protein interactions rather than signaling molecules. This evidence for reciprocal regulation between DARs and NKA provides a novel control mechanism for both DAR signaling and cellular ion balance.

D1 and D2 dopamine receptor expression is regulated by direct interaction with the chaperone protein calnexin.

As for all proteins, G protein-coupled receptors (GPCRs) undergo synthesis and maturation within the endoplasmic reticulum (ER). The mechanisms involved in the biogenesis and trafficking of GPCRs from the ER to the cell surface are poorly understood, but they may involve interactions with other proteins. We have now identified the ER chaperone protein calnexin as an interacting protein for both D(1) and D(2) dopamine receptors. These protein-protein interactions were confirmed using Western blot analysis and co-immunoprecipitation experiments. To determine the influence of calnexin on receptor expression, we conducted assays in HEK293T cells using a variety of calnexin-modifying conditions. Inhibition of glycosylation either through receptor mutations or treatments with glycosylation inhibitors partially blocks the interactions with calnexin with a resulting decrease in cell surface receptor expression. Confocal fluorescence microscopy reveals the accumulation of D(1)-green fluorescent protein and D(2)-yellow fluorescent protein receptors within internal stores following treatment with calnexin inhibitors. Overexpression of calnexin also results in a marked decrease in both D(1) and D(2) receptor expression. This is likely because of an increase in ER retention because confocal microscopy revealed intracellular clustering of dopamine receptors that were co-localized with an ER marker protein. Additionally, we show that calnexin interacts with the receptors via two distinct mechanisms, glycan-dependent and glycan-independent, which may underlie the multiple effects (ER retention and surface trafficking) of calnexin on receptor expression. Our data suggest that optimal receptor-calnexin interactions critically regulate D(1) and D(2) receptor trafficking and expression at the cell surface, a mechanism likely to be of importance for many GPCRs.

The D1 dopamine receptor is constitutively phosphorylated by G protein-coupled receptor kinase 4.

G protein-coupled receptor (GPCR) kinases (GRKs) phosphorylate agonist-activated GPCRs, initiating their homologous desensitization. In this article, we present data showing that GRK4 constitutively phosphorylates the D1 receptor in the absence of agonist activation. This constitutive phosphorylation is mediated exclusively by the alpha isoform of GRK4; the beta, gamma, and delta isoforms are ineffective in this regard. Mutational analysis reveals that the constitutive phosphorylation mediated by GRK4alpha is restricted to the distal region of the carboxyl terminus of the receptor, specifically to residues Thr428 and Ser431. Phosphorylation of the D1 receptor by GRK4alpha results in a decrease in cAMP accumulation, an increase in receptor internalization, and a decrease in total receptor number--all of which are abolished in a D1 receptor mutant containing T428V and S431A. The increase in internalized D1 receptors induced by GRK4alpha phosphorylation is due to enhanced receptor internalization rather than retarded trafficking of newly synthesized receptors to the cell surface. The constitutive phosphorylation of the D1 receptor by GRK4alpha does not alter agonist-induced desensitization of the receptor because dopamine pretreatment produced a similar decrease in cAMP accumulation in control cells versus cells expressing GRK4alpha. These observations shift the attenuation of D1 receptor signaling from a purely agonist-driven process to one that is additionally modulated by the complement of kinases that are coexpressed in the same cell. Furthermore, our data provide direct evidence that, in contrast to current dogma, GRKs can (at least in some instances) constitutively phosphorylate GPCRs in the absence of agonist activation resulting in constitutive desensitization.

The role of phosphorylation in D1 dopamine receptor desensitization: evidence for a novel mechanism of arrestin association.

Homologous desensitization of D(1) dopamine receptors is thought to occur through their phosphorylation leading to arrestin association which interdicts G protein coupling. In order to identify the relevant domains of receptor phosphorylation, and to determine how this leads to arrestin association, we created a series of mutated D(1) receptor constructs. In one mutant, all of the serine/threonine residues within the 3rd cytoplasmic domain were altered (3rdTOT). A second construct was created in which only three of these serines (serines 256, 258, and 259) were mutated (3rd234). We also created four truncation mutants of the carboxyl terminus (T347, T369, T394, and T404). All of these constructs were comparable with the wild-type receptor with respect to expression and adenylyl cyclase activation. In contrast, both of the 3rd loop mutants exhibited attenuated agonist-induced receptor phosphorylation that was correlated with an impaired desensitization response. Sequential truncation of the carboxyl terminus of the receptor resulted in a sequential loss of agonist-induced phosphorylation. No phosphorylation was observed with the most severely truncated T347 mutant. Surprisingly, all of the truncated receptors exhibited normal desensitization. The ability of the receptor constructs to promote arrestin association was evaluated using arrestin-green fluorescent protein translocation assays and confocal fluorescence microscopy. The 3rd234 mutant receptor was impaired in its ability to induce arrrestin translocation, whereas the T347 mutant was comparable with wild type. Our data suggest a model in which arrestin directly associates with the activated 3rd cytoplasmic domain in an agonist-dependent fashion; however, under basal conditions, this is sterically prevented by the carboxyl terminus of the receptor. Receptor activation promotes the sequential phosphorylation of residues, first within the carboxyl terminus and then the 3rd cytoplasmic loop, thereby dissociating these domains and allowing arrestin to bind to the activated 3rd loop. Thus, the role of receptor phosphorylation is to allow access of arrestin to its receptor binding domain rather than to create an arrestin binding site per se.

Melanoma maligno en un quiste dermoide de ovario / Malignant melanoma in a ovarian dermoyd cyst

Presentamos el caso de una paciente a la cual durante una cesárea por sufrimiento fetal agudo se le encuentra una coloración oscura del epiplón, informada posteriormente como melanosis de epiplón altamente sugerente de melanoma. Al examen físico se le detecta una masa hipogástrica que compromete el ovario izquierdo realizándose una ooforectomía izquierda, cuyo resultado fue informado como quiste dermoide con degeneración maligna (melanoma)

Tumor de Ebrikossoff perianal / Perianal Abrikossoff's tumor

Se comunica el caso clínico de una mujer de 57 años que consulta por masa perianal de dos años de evolución, de crecimiento lento. Al exámen físico, masa perianal en sector anterior derecho, de 4 x 3 cm. firme, adherida a planos superficiales y profundos, especialmente al esfínter anal. Biopsia incisional revela tumor benigno decéculas granulares. Se efctúa resección local amplia con excisión parcial de esfínter y sutura esfinteriana. La histología revela un tumor benigno de células granulares o tumor de Abrikossoff. Estudio inmunohistoquímico: Enolasa(++) y proteínas S/100(+) apoya el orígen neural del tumor. Este raro tumor, de histogénesis incierta, que puede presentarse en cualquier sitio del organismo, especilamente en cabeza y cuello(lengua), pared torácica y brazos. Es importante diferenciarlo de la variedad maligna del tumor de células granulares y de los carcinomas de células escamosas, por sus implicancias terapéuticas y pronósticas