ItemArtigoUsing umbilical cord blood as a source of paediatric packed red blood cells: Processing and quality control(2023-01-01) Risso, Mariane Aparecida; Deffune, Elenice [UNESP]; Luzo, Ângela Cristina Malheiros; Universidade Estadual de Campinas (UNICAMP); Universidade Estadual Paulista (UNESP)Background and Objectives: Umbilical cord blood (UCB) has been used as a source of red blood cells (RBCs) for neonatal/paediatric transfusion purposes. This study adopted two different procedures to obtain umbilical RBC (U-RBC) to compare its quality control parameters to those of fractionated adult RBC (A-RBC), for paediatric purposes. Materials and Methods: UCB units (24) were filtered and processed based on two different methods, namely, conventional/manual (P1;n12) and automatic (P2;n12). They were compared to five fractionated A-RBCs. U-RBC and A-RBC were stored for 14 days and had their haematological, biochemical, haemolytic and microbiological parameters analysed at D1, D7 and D14. Cytokines and growth factors (GFs) in residual U-RBC plasma were measured. Results: Mean volume of processed U-RBC units was 45 mL for P1 and 39 mL for P2; the mean haematocrit level reached 57% for P1 and 59% for P2. A-RBC recorded a mean volume of 44 mL. Haematologic and biochemical parameters analysed in U-RBC and A-RBC presented similar behaviours during storage time, except for parameter values, which differed between them. Pro-inflammatory and immunomodulatory cytokines, as well as GFs, were higher in U-RBC residual plasma than in that A-RBC. Conclusion: UCB can be processed into RBC based on either manual or automated protocols. U-RBC units met the referenced quality parameters defined for A-RBC. Some features, mainly the biochemical ones, should be further investigated to help improve quality parameters, with emphasis on differences found in, and particularities of, this material and on recipients of this new transfusion practice. ItemArtigoSiloxane-modified bacterial cellulose as a promising platform for cell culture(2022-12-01) Claro, Amanda Maria; Do Amaral, Nayara Cavichiolli; Colturato, Vitória Maria Medalha; Aleixo, Nadia Andrade; Paiva, Robert; Cruz, Sandra Andrea; Monteiro, Gustavo Claro; De Carvalho, Gustavo Senra Gonçalves; Nogueira, Flávia Aparecida Resende; Deffune, Elenice [UNESP]; Iemma, Mônica Rosas da Costa; Barud, Hernane da Silva; University of Araraquara - UNIARA; Universidade Federal de São Carlos (UFSCar); TechMiP Análises e Soluções Inteligentes LTDA; Universidade Estadual Paulista (UNESP)Cellulose is a versatile tunable material that finds application in the biomedical field as substrate for cell culture. However, few studies have been done to explore the use of bacterial cellulose (BC), a naturally occurring nanofibrillar material with distinct properties, as a platform to produce such device. In the present work, BC membranes have been functionalized with thiol functional group (SH) through silanization reaction with (3-mercaptopropyl)trimethoxysilane (MPTMS) under different conditions, in order to obtain a platform with improved cell adhesion. The efficiency of BC surface modification with MPTMS was evaluated by using acid and base catalyzed reactions, and two different drying methods: at room temperature of 28 °C and at 120 °C as curing temperature. The results of the set of analyses performed—ATR-FTIR, TGA, elemental analysis,13C NMR, contact angle and SEM—indicate that BC surface functionalization was efficient, regardless the drying process. The MPTMS-modified platforms exhibited sulfur content of 3–5 times higher than native BC. The performed biological assay with fibroblast GM07492 human cells revealed that adhesion of cells to the BC surface depends not only on the functional group present at the matrix but also on surface wettability. Graphical abstract: [Figure not available: see fulltext.]Schematic illustration of a siloxane-modified bacterial cellulose as platform for cell culture. ItemArtigoFat grafting: Lipofragmentation X Liposuction(2019-01-01) Mendes, Flavio Henrique [UNESP]; Viterbo, Fausto [UNESP]; Deffune, Elenice [UNESP]; Domingues, Maria Aparecida Custódio [UNESP]; Golim, Marjorie Assis [UNESP]; Gabas, José Marcos [UNESP]; Rossoni, Renan Roldi [UNESP]; Nunes, Helga Caputo [UNESP]; Universidade Estadual Paulista (UNESP)Introduction: Aiming to obtain autogenous and injectable lipografts from resected tissues in dermolipectomies, this study proposes a new method for harvesting and processing adipose tissue through a specific fragmenting device. The main objective was to establish a comparative analysis of the quality and viability characteristics of the new lipofragmentation technique and those of the well-known liposuction technique, widely accepted as a viable source of fat grafting. In vivo and in vitro assays were designed to evaluate the biological behavior of the samples to guide new and possible human studies with clinical applications. Methods: A post-bariatric patient who underwent abdominal dermolipectomy had her surgical specimen resected, which was divided into four parts that underwent liposuction and lipofragmentation, with and without prior infiltration. All samples were centrifuged and distributed for assays with assessments involving histological analysis, immunohistochemistry, flow cytometry, cell culture, and xenograft injection on the back of 10 Wistar rats, which was evaluated after six weeks for mass, volume, and histological features. Results: The structural characteristics and biological behaviors of fragmented, dry, and infiltrated fat samples were similar to those of liposuction samples. Conclusions: Fat fragmentation transformed the subcutaneous cellular tissue of dermolipectomies into a new, viable injectable lipograft variant, with biological characteristics similar to those of traditional liposuction. Although still preliminary, our results support further investigations to optimize the technique and improve fat grafting and its possible applications in regenerative medicine. ItemArtigoProteoglycans orchestrate remodeling of prostatic cytoarchitecture after androgenic blockade in old gerbils(2023-02-01) Campos, Silvana G. P. [UNESP]; Gonçalves, Bianca F. [UNESP]; Ruiz, Thalles Fernando Rocha; Leonel, Ellen Cristina R.; Ribeiro, Daniele L.; Falleiros Junior, Luiz Roberto; Goes, Rejane M. [UNESP]; Taboga, Sebastião R. [UNESP]; Universidade Estadual Paulista (UNESP); Universidade Estadual de Campinas (UNICAMP); Universidade Federal de Goiás (UFG); Universidade Federal de Uberlândia (UFU); São José Rio Preto School of Medicine (FAMERP)Background: The aim of this study was to evaluate modifications in proteoglycan morphology and composition in the prostatic stroma of 18-month-old gerbils after surgical castration, in association or not with an androgenic blockade. Methods: The animals (n = 5) were sorted into groups subjected or not to antiandrogen treatment (flutamide 10 mg/kg/day) administered for the total postsurgery period and euthanized at 7- or 30-day postcastration; the control group consisted of intact animals. Tissue analysis included immunohistochemical assessment (perlecan and chondroitin sulfate) and proteoglycan morphology was analyzed by transmission electron microscopy. Results: Chondroitin sulfate frequency was increased 7 days postcastration with an androgenic blockade. The presence of these carbohydrates was rare after 30 days of androgenic blockade treatment. There was a significant increase in the amount of perlecan in the prostate stroma from groups subjected to castration plus flutamide for 7 or 30 days. Ultrastructural analysis showed that the incidence of areas occupied by proteoglycans and basement membrane was altered by treatment. In addition, androgenic blockade results in changes in the amount, thickness, and morphology of these structures. At 30 days postcastration, with or without flutamide treatment, larger proteoglycans were common. Conclusions: In this study, in particular, the decrease in chondroitin sulfate after the longer period might be understood as a prostatic response to androgenic deprivation, while the high frequency and permanence of perlecan led to the assumption that its modulation could be androgen-independent. Length and form alterations in proteoglycans as well as associations among them and with the basement membrane were dynamic events in the prostate microenvironment. ItemArtigoHow I Do It: Transperineal prostate biopsy using local anesthetic in an outpatient setting(Canadian J Urology, 2023-02-01) Ordones, Flavio Vasconcelos [UNESP]; Vermeulen, Lodewikus; Bressington, Morgan; Menon, Abilash; Burns, Timothy; Muller, Loretta; Fraundorfer, Mark; Gilling, Peter J.; Tauranga Publ Hosp; Universidade Estadual Paulista (UNESP); Univ AucklandTransperineal prostate biopsy (TPPB) is proven to be an effective diagnostic tool for prostate cancer detection. It allows satisfactory sampling of apical and anterior areas which is not well achieved with the transrectal route, without the associated risks of urinary tract infection or sepsis. The main objective of this paper is to describe the technique utilized in our institution to perform transperineal prostate biopsy under local anesthetic in the outpatient clinic setting. ItemArtigoBisphenol A and 2,3,7,8-tetrachlorodibenzo-p-dioxin at non-cytotoxic doses alter the differentiation potential and cell function of rat adipose-stem cells(2022-09-01) Nunes, Helga Caputo [UNESP]; Tavares, Samara Costa [UNESP]; Garcia, Heloísa Vicente [UNESP]; Cucielo, Maira Smaniotto [UNESP]; dos Santos, Sérgio Alexandre Alcântara [UNESP]; Aal, Mirian Carolini Esgoti [UNESP]; de Golim, Marjorie Assis [UNESP]; Justulin, Luís Antônio [UNESP]; Ribeiro, Amanda Oliveira [UNESP]; Deffune, Elenice [UNESP]; Scarano, Wellerson Rodrigo [UNESP]; Delella, Flávia Karina [UNESP]; Universidade Estadual Paulista (UNESP)The possibility of chemical contamination is an important issue to consider when designing a cell therapy strategy. Both bisphenol A (BPA) and 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) are among the most environmentally relevant endocrine disrupting chemicals (EDCs, compounds with a high affinity for adipose tissue) recently studied. Adipose-derived stem cells (ASCs) are mesenchymal stromal cells (MSCs) obtained from adipose tissue widely used in regenerative medicine to prevent and treat diseases in several tissues and organs. Although the experimental use of tissue-engineered constructs requires careful analysis for approval and implantation, there has been a recent increase in the number of approved clinical trials for this promising strategy. This study aimed to evaluate cell viability, apoptosis, DNA damage, and the adipogenic or osteogenic differentiation potential of rat adipose-derived stem cells (rASCs) exposed to previously established non-cytotoxic doses of BPA and TCDD in vitro. Results demonstrated that 10 μM of BPA and 10 nM of TCDD were able to significantly reduce cell viability, while all exposure levels resulted in DNA damage, although did not increase the apoptosis rate. According to the analysis of adipogenic differentiation, 1 μM of BPA induced the significant formation of oil droplets, suggesting an increased adipocyte differentiation, while both 10 μM of BPA and 10 nM of TCDD decreased adipocyte differentiation. Osteogenic differentiation did not differ among the treatments. As such, BPA and TCDD in the concentrations tested can modify important processes in rASCs such as cell viability, adipogenic differentiation, and DNA damage. Together, these findings prove that EDCs play an important role as contaminants, putatively interfering in cell differentiation and thus impairing the therapeutic use of ASCs. ItemArtigoAlzheimer's disease diagnosis based on detection of autoantibodies against Aβ using Aβ40 peptide in liposomes(2022-06-01) Monteiro, Júlio César; Yokomichi, Anna Laura Yuri; de Carvalho Bovolato, Ana Lívia [UNESP]; Schelp, Arthur Oscar [UNESP]; Ribeiro, Sidney José Lima [UNESP]; Deffune, Elenice [UNESP]; Moraes, Marli Leite de; Universidade Federal de São Paulo (UNIFESP); Universidade Estadual Paulista (UNESP)Background: Alzheimer's disease (AD) is the most common form of dementia and affect more than 50 million people worldwide. Thus, there is a high demand by non-invasive methods for an early diagnosis. This work explores the AD diagnostic using the amyloid beta 1–40 (Aβ40) peptide encapsulated into dipalmitoyl phosphatidyl glycerol (DPPG) liposomes and immobilized on polyethylene imine previously deposited on screen-printed carbon electrodes to detect autoantibodies against Aβ40, a potential biomarker found in plasma samples. Methods: The immunosensor assembly was accompanied by atomic force microscopy (AFM) images that showed globular aggregates from 20 to 200 nm corresponding liposomes and by cyclic voltammetry (CV) through increase of the voltammogram area each material deposited. After building the immunosensor, when it was exposed to antibody anti-Aβ40, there was an increase in film roughness of approximately 9 nm, indicating the formation of the immunocomplex. Results: In the detection by CV, the presence of specific antibody, in the range of 0.1 to 10 μg/ml, resulted in an increase in the voltammograms area and current in 0.45 V reaching 3.2 µA.V and 5.7 μA, respectively, in comparison with the control system, which remained almost unchanged from 0.1 μg/ml. In patient samples, both cerebrospinal fluid (CSF) and plasma, was possible separated among positive and negative samples for AD using CV profile and area, with a difference of 0.1 μA.V from the upper error bar of healthy samples for CSF sample and 0.6 μA.V for plasma sample. Conclusions: These results showed the feasibility of the method employed for the non-invasive diagnostic of Alzheimer's disease detecting natural autoantibodies that circulate in plasma through a simple and easy-to-interpret method. ItemArtigoFasting and 24-h urine pH in patients with urolithiasis using potassium citrate(E-century Publishing Corp, 2022-01-01) Silva, Isabela Bertanholi Leme da [UNESP]; Amarol, Carmen Petean [UNESP]; Amaro, Joao Luiz [UNESP]; Cunha, Natalia Baraldi [UNESP]; Callegari, Matheus Augusto [UNESP]; Yamamoto, Hamilto Akihissa [UNESP]; Guerra, Rodrigo [UNESP]; Quitzan, Juliany Gomes [UNESP]; Reis, Leonardo O.; Kawano, Paulo Roberto [UNESP]; Universidade Estadual Paulista (UNESP); Universidade Estadual de Campinas (UNICAMP)Purpose: To evaluate the correlation between the pH readings in 24 h urine and the random fasting specimen in patients with urolithiasis using 2 methods. Methods: A total of 114 patients with urinary lithiasis using potassium citrate were prospectively analyzed. All patients collected 24-h urine and an additional sample, after nocturnal fasting, collected on the day they brought the 24-h sample at the lab. Two different methods (test strip and digital meter) were used to determine pH values. Results: The pH analysis using strips in the 24-h urine presented a mean value similar to the one obtained in the fasting sample (6.07 +/- 0.74 vs. 6.02 +/- 0.82, respectively; P > 0.05). The same behavior was seen considering the readings with a digital pH meter (5.8 +/- 0.78 vs. 5.75 +/- 0.83; P > 0.05). However, readings conducted in the same specimen with pH meter and test strip were dissonant (P < 0.05), suggesting that the colorimetric method is not reliable in the assessment of urinary pH in this population. Conclusion: pH assessment in a random urinary specimen proved as efficient as the 24-h urine standard method to monitor patients with kidney stones in the use of potassium citrate. Classical test strip analysis is not sensitive enough to evaluate the urine pH in this population and digital pH meter reading is preferred. ItemResenhaAcute graft thrombosis in patients who underwent renal transplant and received anticoagulant or antiplatelet agents. A systematic review and meta-analysis(E-century Publishing Corp, 2022-01-01) Guerra, Rodrigo [UNESP]; Kawano, Paulo Roberto [UNESP]; Amaro, Marcelo Petean; Yamamoto, Hamilto Akihissa [UNESP]; Gomes Filho, Fernando Ferreira [UNESP]; Amaro, Joao Luiz [UNESP]; El Dibl, Regina Paolucci [UNESP]; Garcia-Perdomo, Herney Andres; Reis, Leonardo Oliveira; Universidade Estadual Paulista (UNESP); Univ Valle; Universidade Estadual de Campinas (UNICAMP)Objectives: Thrombosis is a major cause of early allograft loss in renal transplantation. Herein, we assessed the frequency of acute graft thrombosis in patients who underwent renal transplant and received anticoagulant or antiplatelet agents. Methods: We performed a systematic review of all available case series studies of anticoagulant and/or antiplatelet prophylaxis of thrombosis in renal transplantation. The data were pooled in a proportional meta-analysis. Results: Twenty-one case series were identified from 7,160 retrieved titles. A total of 3,246 patients were analyzed (1,718 treated with antiplatelet and/or anticoagulant agents and 1,528 non-treated control subjects). Allograft thrombosis occurred in 7.24% (95% CI 3.45 to 12.27%) of the patients receiving no intervention compared with 3.38% (95% CI 1.45 to 6.1%), 1.2% (95% CI 0.6 to 2.1%) and 0.47% (95% CI 0.001 to 1.79%) of the patients in the anticoagulant, aspirin, and aspirin + anticoagulant groups, respectively. The bleeding complication rate for anticoagulants was significantly higher than in the other groups. Conclusions: Our data suggests that anticoagulants, and aspirin, either alone or in association with an anticoagulant, seem to have a low frequency of acute allograft thrombosis after kidney transplantation. Higher hemorrhagic complication rates might occur when anticoagulants are used. ItemResumo3-D Printed prostate cancer model prior to Robotic-Assisted Radical Prostatectomy: Initial experience and a surgeon, patient, radiologist, and pathologist point of view(Elsevier B.V., 2022-02-01) Ordones, Vasconcelos F.; Araujo, L. F. Takano de [UNESP]; Fornazieri, L. F.; Artioli, B. A. Olivetti [UNESP]; Kawano, P. K. Roberto [UNESP]; Amaro, J. L. [UNESP]; Leite, K. R.; Tauranga Hosp; Universidade Estadual Paulista (UNESP); AC Camargo Canc Ctr; Genoa Biotech ItemArtigoPenicillin-binding proteins (PBPs) determine antibiotic action in Langmuir monolayers as nanoarchitectonics mimetic membranes of methicillin-resistant Staphylococcus aureus(2022-06-01) Martins, Beatriz Araújo; Deffune, Elenice [UNESP]; Oliveira, Osvaldo N.; Moraes, Marli Leite de; Universidade de São Paulo (USP); Universidade Estadual Paulista (UNESP)The membrane of methicillin-resistant Staphylococcus aureus (MRSA) contains penicillin-binding proteins (PBPs) in the phospholipidic bilayer, with the protein PBP2a being linked with the resistance mechanism. In this work we confirm the role of PBP2a with molecular-level information obtained with Langmuir monolayers as cell membrane models. The MRSA cell membrane was mimicked with a mixed monolayer of dipalmitoyl phosphatidyl glycerol (DPPG) and cardiolipin (CL), also incorporating PBP2a. The surface pressure-area isotherms and the Brewster angle microscopy (BAM) images for these mixed monolayers were significantly affected by the antibiotic meropenem, which is PBP2a inhibitor. The meropenem effects were associated with the presence of PBP2a, as they were absent in the Langmuir monolayers without PBP2a. The relevance of PBP2a was confirmed with results where the antibiotic methicillin, known to be unsuitable to kill MRSA, had the same effects on mixed DPPG/CL and DPPG/CL-PBP2a monolayers since it prevented PBP2a from incorporating in the monolayer. The biological implication of the findings presented here is that a successful antibiotic against MRSA should be able to interact with PBP2a, but in the membrane. ItemArtigoScreen-Printed Electrodes on Tyvek Substrate as Low-Cost Device to Applications in Alzheimer's Disease Detection(2022-03-01) Palley, Bianca Fortes; Artur, Julio Cesar; De Arruda, Milena Nakagawa; De Souza, Gustavo Freitas; Graves, David Alexandro; Bovolato, Ana Lívia De Carvalho [UNESP]; Deffune, Elenice [UNESP]; Schelp, Arthur Oscar [UNESP]; Gonçalves, Emerson Sarmento; De Moraes, Marli Leite; Instituto Tecnológico de Aeronaútica; Universidade Federal de São Paulo (UNIFESP); Universidade Do Vale Do Paraíba; Instituto de Aeronáutica e Espaço; Universidade Estadual Paulista (UNESP)Alzheimer's disease (AD) is a progressive neurodegenerative disease characterized by impairment of cognitive functions and memory deterioration, which requires an early diagnosis for effective treatment. The use of immunosensors to detect Alzheimer s disease biomarkers stand out as a quick and cheap alternative for early detection of the disease. The development of Screen-Printed Electrodes (SPEs) meets a growing demand in the market for applications such as signal transducers in biosensor devices. In this work was development a method for fabrication of low cost SPE for application in the detection of Alzheimer s disease through autoantibodies. It was produced microelectrodes on polyvinyl chloride (PVC), polyvinylidene fluoride (PVDF) and high-density polyethylene (HDPE) manufactured by DuPont Tyvek®substrates. SPEs produced on Tyvek®substrates have shown promising results for low-cost, disposable and flexible sensors. The carbon paste showed excellent adhesion to Tyvek®substrate and the electrodes produced showed an electrochemical performance comparable to commercial electrodes, besides reproducible. The detection results showed that it is possible to detect anti-Aβ40 autoantibodies in real samples of serum and cerebrospinal fluid using the electrode modified with two bilayers of (PEI/(DDPG + Aβ40)) since the capacitance increased more in positive sample with the presence of autoantibody than in negative samples. ItemArtigoAnti-aquaporin-4 immunoglobulin G colorimetric detection by silver nanoparticles(2022-04-01) Higa, Akemi M.; Moraes, Ariana S.; Shimizu, Flávio M.; Bueno, Raquel G.; Peroni, Luís A.; Strixino, Francisco T.; Sousa, Nise A.C.; Deffune, Elenice [UNESP]; Bovolato, Ana Lívia C. [UNESP]; Oliveira, Osvaldo N.; Brum, Doralina G. [UNESP]; Leite, Fabio L.; Universidade de São Paulo (USP); Universidade Federal de São Carlos (UFSCar); Brazilian Center for Research in Energy and Materials; Rheabiotech Laboratory of Research and Development; Universidade Federal do Amazonas; Universidade Estadual Paulista (UNESP)Neuromyelitis optica spectrum disorder (NMOSD) is an inflammatory and autoimmune disease whose biomarker is the anti-AQP4-IgG autoantibody that binds to aquaporin-4 (AQP4) protein. We introduced a nanosensor with a sensitivity of 84.6%, higher than the CBA's 76.5%. Using silver nanoparticles (AgNPs), we detected not only seropositive but also some false-negative patients previously classified with CBA. It consisted of AgNPs coated with one of a panel of 5 AQP4 epitopes. The ability in detecting the anti-AQP4-IgG in NMOSD patients depended on the epitope and synergy could be obtained by combining different epitopes. We demonstrated that NMOSD patients could easily be distinguished from healthy subjects and patients with multiple sclerosis. Using the most sensitive AQP461-70 peptide, we established a calibration curve to estimate the concentration of anti-AQP4-IgG in seropositive NMOSD patients. The ability to enhance the accuracy of the diagnosis may improve the prognosis of 10-27% of anti-AQP4-IgG seronegative patients worldwide. ItemArtigoProbeless and label-free impedimetric biosensing of D-dimer using gold nanoparticles conjugated with dihexadecylphosphate on screen-printed carbon electrodes(2021-11-20) Tasić, Nikola; Cavalcante, Letícia; Deffune, Elenice [UNESP]; Góes, Márcio Sousa; Paixão, Thiago R.L.C.; Gonçalves, Luís Moreira; Universidade de São Paulo (USP); Universidade Estadual Paulista (UNESP); Universidade Federal da Integração Latino-Americana (UNILA)D-dimer (DD) is a clinical biomarker of emerging significance. Its fast analysis and quantification, preferably by the bedside, helps a medical doctor make crucial decisions. Electrochemical biosensors are point-of-care technologies that can address such issues. Herein, it is developed a probeless and label-free impedimetric DD biosensor. Gold nanoparticles (AuNPs) are dispersed along with dihexadecylphosphate (DHP) on the surface of screen-printed carbon electrodes (SPCEs) to attach anti-DD monoclonal antibody as sensing recognition element, and the measurements are performed using electrochemical impedance spectroscopy (EIS). Two different analytical models are used to interpret raw impedance spectra. The first model is based on the capacitive response within the DHP monolayer and its electrochemical occupancy described by the complex capacitance value at the frequency of 200 mHz, noted as M1. The second model is based on the charge transfer resistance changes (Rct) occurring upon the mAb-DD binding event, fitted by two different Randles types of the equivalent circuits (noted as M2a and M2b). The models are compared, and with the obtained high linearity in the clinically relevant range (up to 500 ng mL−1) and low levels of detection (as low as 8.92 ng mL−1), a potential POC sensor is demonstrated. Additionally, the proposed sensor is applied in real whole blood samples showing good recovery values with certain discrepancies from the standard laboratory assay result, making a cheap and straightforward way for semi-quantitative bedside clinical evaluation. ItemArtigoXeroderma pigmentosum: Low prevalence of germline XPA mutations in a brazilian XP population(2015-04-01) Santiago, Karina Miranda; De Nóbrega, Amanda França; Rocha, Rafael Malagoli; Rogatto, Silvia Regina; Achatz, Maria Isabel; A.C. Camargo Cancer Center; National Institute of Science and Technology in Oncogenomics (INCITO); Universidade de São Paulo (USP)Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder characterized by DNA repair defects that cause photophobia, sunlight-induced cancers, and neurodegeneration. Prevalence of germline mutations in the nucleotide excision repair gene XPA vary significantly in different populations. No Brazilian patients have been reported to carry a germline mutation in this gene. In this study, the germline mutational status of XPA was determined in Brazilian patients exhibiting major clinical features of XP syndrome. The study was conducted on 27 unrelated patients from select Brazilian families. A biallelic inactivating transition mutation c.619C>T (p.Arg207Ter) was identified in only one patient with a history of neurological impairment and mild skin abnormalities. These findings suggest that XP syndrome is rarely associated with inherited disease-causing XPA mutations in the Brazilian population. Additionally, this report demonstrates the effectiveness of genotype-phenotype correlation as a valuable tool to guide direct genetic screening. ItemArtigoDevelopment of a rabbit's urethral sphincter deficiency animal model for anatomical-functional evaluation(2012-08-30) Skaff, M.; Pinto, E. R.S.; Leite, K. R.M.; Almeida, F. G.; Universidade de São Paulo (USP)Objective: The aim of the study was to develop a new durable animal model (using rabbits) for anatomical-functional evaluation of urethral sphincter deficiency. Materials and Methods: A total of 40 new Zealand male rabbits, weighting 2.500 kg to 3.100 kg, were evaluated to develop an incontinent animal model. Thirty-two animals underwent urethrolysis and 8 animals received sham operation. Before and at 2, 4, 8 and 12 weeks after urethrolysis or sham operation, it was performed cystometry and leak point pressure (LPP) evaluation with different bladder distension volumes (10, 20, 30 mL). In each time point, 10 animals (8 from the study group and 2 from the sham group) were sacrificed to harvest the bladder and urethra. The samples were evaluated by H & E and Masson's Trichrome to determine urethral morphology and collagen/smooth muscle density. Results: Twelve weeks after urethrolysis, it was observed a significant decrease in LPP regardless the bladder volume (from 33.7 ± 6.6 to 12.8 ± 2.2 cmH2O). The histological analysis evidenced a decrease of 22% in smooth muscle density with a proportional increase in the collagen, vessels and elastin density (p < 0.01). Conclusions: Transabdominal urethrolysis develops urethral sphincter insufficiency in rabbits, with significant decrease in LPP associated with decrease of smooth muscle fibers and increase of collagen density. This animal model can be used to test autologous cell therapy for stress urinary incontinence treatment. ItemArtigoBladder augmentation in rabbits with anionic collagen membrane, with or without urotelial preservation. Cistometric and hystologic evaluation(2002-09-01) Lepper, Fábio Gustavo Oliveira; Ramos, Tamara M.; Trindade Filho, José C.S.; Vale, Fabiana R.; Padovani, Carlos R.; Goissis, Gilberto; Universidade de São Paulo (USP)Introduction: The use of bowel segments to perform bladder augmentation is associated with several metabolic and surgical complications. A great variety of synthetic materials, biodegradable or not, have been tested. Collagen-based biomaterials have shown effectiveness for the regeneration and obtainment of a functional bladder. Objective: Assess the functional and histological response of the rabbit bladder to anionic collagen membrane (ACM), either when it is anastomosed to the bladder or it is placed onto bladder after vesicomyectomy. Materials and methods: In 15 male rabbit a partial cystectomy was performed. After 4 weeks they were divided in 3 groups. Group 1 (G1)-bladder augmentation with ACM. Group 2 (G2) ACM is placed onto bladder after vesicomyectomy. Group 3 (G3) control group. Maximal bladder capacity (MBC) and weight were assessed with 4 (M1), 8 (M2) and 12 (M3) weeks after partial cystectomy. In M3 was performed the sacrifice and extraction of the bladder and kidneys for anatomopathologic study. Results: There were neither bladder stones, nor implant extrusion in M3. There was a significant increase in MBC in G1 and G2 (p<0.05), but no statistical differences in G3 (p=0.35). There is no significant difference comparing G1 and G2. In M3, both groups have shown a bigger MBC than G3 (p<0.05). The microscopic assessment showed an inflammatory reaction in the bladder augmented, with urothelium preserved. Conclusions: The ACM was effective for the increase of MBC. The bladders with preservation of the urothelium have shown an extensive inflammatory process. ItemArtigoMultiple analyses suggests texture features can indicate the presence of tumor in the prostate tissue(2022-01-01) Souza, Sérgio Augusto Santana [UNESP]; Reis, Leonardo Oliveira; Alves, Allan Felipe Fattori [UNESP]; Silva, Letícia Cotinguiba [UNESP]; Medeiros, Maria Clara Korndorfer; Andrade, Danilo Leite; Billis, Athanase; Amaro, João Luiz [UNESP]; Martins, Daniel Lahan; Trindade, André Petean [UNESP]; Miranda, José Ricardo Arruda [UNESP]; Pina, Diana Rodrigues [UNESP]; Universidade Estadual Paulista (UNESP); Universidade Estadual de Campinas (UNICAMP)Several studies have demonstrated statistical and texture analysis abilities to differentiate cancerous from healthy tissue in magnetic resonance imaging. This study developed a method based on texture analysis and machine learning to differentiate prostate findings. Forty-eight male patients with PI-RADS classification and subsequent radical prostatectomy histopathological analysis were used as gold standard. Experienced radiologists delimited the regions of interest in magnetic resonance images. Six different groups of images were used to perform multiple analyses (seven analyses variations). Those analyses were outlined by specialists in urology as those of most significant importance for the classification. Forty texture features were extracted from each image and processed with Random Forest, Support Vector Machine, K-Nearest Neighbors, and Naive Bayes. Those seven analyses variation results were described in terms of area under the ROC curve (AUC), accuracy, F-score, precision and sensitivity. The highest AUC (93.7%) and accuracy (88.8%) were obtained when differentiating the group with both MRI and histopathology positive findings against the group with both negative MRI and histopathology. When differentiating the group with both MRI and histopathology positive findings versus the peripheral image zone group the AUC value was 86.6%. When differentiating the group with negative MRI/positive histopathology versus the group with both negative MRI and histopathology the AUC value was 80.7%. The evaluation of statistical and texture analysis promoted very suggestive indications for future work in prostate cancer suspicious regions. The method is fast for both region of interest selection and classification with machine learning and the result brings original contributions in the classification of different groups of patients. This tool is low-cost, and can be used to assist diagnostic decisions. ItemArtigoMultiple Tolerization Subtractive Immunization (MTSI) Protocol: Effects on Mice and Monoclonal Antibody Specificity(2021-12-07) de Lima Fontes, Marina [UNESP]; Neves, Franciny Mara de Lima [UNESP]; Santos, Kelvin Sousa [UNESP]; Fusco-Almeida, Ana Marisa [UNESP]; Giannini, Maria José Soares Mendes [UNESP]; Felisbino, Sergio Luis [UNESP]; Deffune, Elenice [UNESP]; Moroz, Andrei [UNESP]; Universidade Estadual Paulista (UNESP)Monoclonal antibodies (mAbs) have been a valuable tool to elucidate several biological processes, such as stem cell differentiation and cancer, and contributed to virtually all areas of biomedical sciences. Yet, it remains a challenge to obtain mAbs specific to poorly expressed epitopes, or to epitopes that are actually involved in important biological phenomena, such as cell differentiation and metastasis. Drug-induced subtractive immunization, and recently the multiple tolerization subtractive immunization (MTSI) technique, reported by our group, have the potential to level up the field, as they direct the host´s immune response towards these epitopes. However, due to cyclophosphamide (CY) treatment, high mice mortality can be observed, and only a few data are available on how these techniques affect the immune system of mice. Tolerogen and immunogen cells, RWPE-1 and PC-3 cells, respectively, were individually seeded at 2 × 104 cells/cm2, and then adjusted to 2 × 106 cells per mouse before immunization, which was conducted in a subtractive approach (MTSI) with CY. Immunosuppression of mice was recorded via total white blood counting, as well the reactivity of circulating polyclonal antibodies (pAbs). General parameters, including weight, physical appearance, and behavior on mice subjected to three different concentrations of CY were recorded. mAbs were obtained using classical hybridoma techniques, using the spleen of immunized mice. After purification, antibodies were characterized by Western blotting, and Indirect immunofluorescence. In conclusion, all CY dosage were efficient in creating an immunosuppression state, but only the 100 mg/kg body weight was feasible, as the others resulted in extensive mice mortality. pAbs obtained in the peripheral blood of mice showed more reactivity towards tumor cells. MAbs 2-7A50 and 2-5C11 recognized antigens from tumor cells, but not from their non-tumor counterparts, as shown in western blotting and immunofluorescence assays. MTSI technique was successful in generating mAbs that recognize tumor-specific antigens. ItemArtigoA comparative analisys of pelvic floor muscle strength in women with stress and urge urinary incontinence(2012-12-31) Gameiro, Monica Orsi [UNESP]; Moreira, Eliane Cristina; Ferrari, Renata Spagnoli [UNESP]; Kawano, Paulo Roberto [UNESP]; Padovani, Carlos Roberto [UNESP]; Amaro, João Luiz [UNESP]; Universidade Estadual Paulista (UNESP); Paraná State UniversityAims: To assess pelvic floor muscle (PFM) strength in women with stress urinary incontinence (SUI) and urge urinary incontinence (UUI). Materials and Methods: 51 women were prospectively divided into two groups, according to the symptoms as SUI (G1 = 22) or UUI (G2 = 29). Demographic data, such as number of pads 24 hours, number of micturations 24 hours and nocturia, delay time of urgent void (i.e., the time period for which an urgent void could be voluntarily postponed), number of parity and vaginal deliveries were obtained using a clinical questionnaire. Objective urine loss was evaluated by 60-min. Pad Test, subjective urine stream interruption test (UST) and visual survey of perineal contraction. Objective evaluations of PFM were performed in all patients (vaginal manometry). Results: Median of age, mean number of pads 24 hours, nocturia and warning time were significantly higher in UUI comparing to SUI group. During UST, 45.45% in G1 and 3.44%, in G2, were able to interrupt the urine stream (p < 0.001). The 60-min. Pad Test was significantly higher in G2 compared to G1 women (2.7 ± 2.4 vs 1.5 ± 1.9 respectively, p = 0.049). Objective evaluation of PFM strength was significantly higher in the SUI than in the UUI patients. No statistical difference was observed regarding other studied parameters. Conclusion: Pelvic floor muscle weakness was significantly higher in women with UUI when compared to SUI.