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PURPOSE: To assess the utility of pig head model as an oculoplastic surgical training tool. METHODS: Fresh pig head was availed for surgical workshop for entropion correction, eyelid margin repair, and evisceration with implant for oculoplastic surgery-naïve participants. Skill improvement, surgical understanding, and performance of the participants were evaluated both subjectively and objectively by trained oculoplastic surgeons. Subjective assessment was done by a standardized questionnaire based on Likert scale shared with the participants post training. Objective evaluation was done by the faculty based on a three-point scale and a competency-based surgical assessment rubric. RESULTS: There were 18 surgery-naïve participants in the workshop which comprised ophthalmology residents and comprehensive ophthalmologists. About 88.88% of the participants were able to perform the lid margin and sub-tarsal dissection in entropion surgery. While performing lid tear repair, 94.44% were correctly able to identify the grey line and anterior and posterior lamellae. About 83.33% of the participants were able to place an implant in the scleral shell or intraconal space. About 83.33% of the participants noted that texture and tissue maneuvering were similar to the human eye while performing surgical steps. About 94.44% of the participants reported better understanding, development of skill and additional confidence after training. The median score on objective assessment was 3. The performance on real patients resulted in a median score of 4 for entropion correction. CONCLUSION: Porcine orbital dissection is an available, affordable, and useful tool for oculoplastic surgical training. Training on porcine model can improve anatomical understanding, clinical judgement, and surgical efficiency in trainees.
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Glycation and the subsequent formation of advanced glycation end products (AGEs) disrupt and impair the physiological functions of proteins. This study presents a comprehensive glycation site mapping of human serum albumin (HSA) utilizing liquid chromatography-tandem mass spectrometry (LC-MS/MS). Both in vitro glycation experiments and patient samples were investigated, exploring various enzymes, processing techniques, and their impacts on glycation site detection. A pilot study was conducted, analyzing sixteen serum samples, which spanned from healthy individuals to severe diabetic patients (with HbA1c values ranging from 5.7% to 18.1%). The aim was to comprehend the progression of glycation on various sites of HSA with increasing levels of glycation. Their glycated albumin levels (GA) spanned from 19.7% to 62.3%. Trypsin-mediated proteolytic digestion unveiled 12 glycation sites through direct in-solution digestion of whole serum. However, isolating albumin from serum enabled the identification of a higher number of glycation sites in each sample compared to direct serum digestion. Boronate affinity chromatography facilitated the segregation of less glycated albumin (LGA) from the more glycated albumin (MGA) fraction. Subsequent proteolytic digestion of both LGA and MGA samples revealed similar glycation sites. The MGA fraction exhibited a greater number of identified glycation sites, thereby elucidating which sites are particularly prone to glycation in highly glycated albumin samples. Changes in relative glycation levels were noted in the tryptic digests of albumin samples following the sample enrichment steps, as opposed to direct in-solution digestion of whole serum. Two enzymes, trypsin and Glu-C, were evaluated for efficacy in sequence coverage and glycation site analysis of HSA, with trypsin demonstrating superior efficiency over Glu-C.
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Albumina Sérica Glicada , Produtos Finais de Glicação Avançada , Albumina Sérica Humana , Espectrometria de Massas em Tandem , Humanos , Produtos Finais de Glicação Avançada/química , Espectrometria de Massas em Tandem/métodos , Albumina Sérica Humana/química , Glicosilação , Cromatografia Líquida/métodos , Albumina Sérica/química , Albumina Sérica/análise , Albumina Sérica/metabolismo , Projetos Piloto , Diabetes Mellitus/sangue , Tripsina/química , Tripsina/metabolismoRESUMO
Aims: Cys34 albumin redox modifications (reversible "cysteinylation" and irreversible "di/trioxidation"), besides being just oxidative stress biomarkers, may have primary pathogenetic roles to initiate and/or aggravate cell, tissue, and vascular damage in diabetes. In an exploratory "proof-of-concept" pilot study, we examined longitudinal changes in albumin oxidation during diabetes therapy. Methods: Mass spectrometric analysis was utilized to monitor changes in human serum albumin (HSA) post-translational modifications {glycation [glycated albumin (GA)], cysteinylation [cysteinylated albumin (CA) or human non-mercaptalbumin-1; reversible], di/trioxidation (di/trioxidized albumin or human non-mercaptalbumin-2; irreversible), and truncation (truncated albumin)} during ongoing therapy. Four informative groups of subjects were evaluated [type 1 diabetes (T1DM), type 2 diabetes (T2DM), prediabetes-obesity, and healthy controls] at baseline, and subjects with diabetes were followed for a period up to 280 days. Results: At baseline, T2DM was associated with relatively enhanced albumin cysteinylation (CA% total) compared with T1DM (P = 0.004), despite comparable mean hyperglycemia (P values: hemoglobin A1c = 0.09; GA = 0.09). T2DM, compared with T1DM, exhibited selectively and significantly higher elevations of all the "individual" glycated cum cysteinylated ("multimodified") albumin isoforms (P values: CysHSA+1G = 0.003; CysHSA+2G = 0.007; and CysHSA+3G = 0.001). Improvements in glycemic control and decreases in albumin glycation during diabetes therapy in T2DM were not always associated with concurrent reductions of albumin cysteinylation, and in some therapeutic situations, albumin cysteinylation worsened (glycation-cysteinylation discordance). Important differences were observed between the effects of sulfonylureas and metformin on albumin molecular modifications. Conclusions: T2DM was associated with higher oxidative (cysteinylation) and combined (cysteinylation plus glycation) albumin molecular modifications, which are not ameliorated by improved glucose control alone. Further studies are required to establish the clinical significance and optimal therapeutic strategies to address oxidative protein damage and resulting consequences in diabetes.
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Diabetes Mellitus Tipo 2 , Albumina Sérica Glicada , Hipoglicemiantes , Oxirredução , Albumina Sérica Humana , Humanos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/metabolismo , Masculino , Pessoa de Meia-Idade , Feminino , Hipoglicemiantes/uso terapêutico , Albumina Sérica Humana/metabolismo , Albumina Sérica Humana/química , Glicosilação , Projetos Piloto , Adulto , Albumina Sérica/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Biomarcadores/sangue , Hemoglobinas Glicadas/metabolismo , Hemoglobinas Glicadas/análise , Glicemia/metabolismo , Estudos de Casos e Controles , Idoso , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/sangue , Diabetes Mellitus Tipo 1/metabolismo , Produtos Finais de Glicação Avançada/metabolismo , Processamento de Proteína Pós-Traducional , Metformina/uso terapêutico , Cisteína/metabolismoRESUMO
Sickle cell disease (SCD) is characterized by hemolysis, vaso-occlusion, and ischemia. HIV-1 infection was previously shown to be suppressed in SCD PBMCs. Here, we report that HIV-1 suppression is attributed to the increased expression of iron, hypoxia, and interferon-induced innate antiviral factors. Inhibition of upregulated antiviral genes, HMOX-1, CDKN1A, and CH25H, increased HIV-1 replication in SCD PBMCs, suggesting their critical role in HIV-1 suppression. Levels of IFN-ß were elevated in SCD patients. Sickle cell hemoglobin (HbS) treatment of THP-1-derived and primary monocyte-derived macrophages induced production of IFN-ß, upregulated antiviral gene expression, and suppressed HIV-1 infection. Infection with mouse-adapted EcoHIV was suppressed in the SCD mice that also exhibited elevated levels of antiviral restriction factors. Our findings suggest that hemolysis and release of HbS leads to the induction of IFN-ß production, induction of cellular antiviral state by the expression of iron and IFN-driven factors, and suppression of HIV-1 infection.