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1.
World J Urol ; 38(9): 2221-2226, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31781895

RESUMO

PURPOSE: To describe the overall extirpative renal surgery (ERS) training volume reported by PU and PS. METHODS: Case log data from the Accreditation Council for Graduate Medical Education (ACGME) was examined from 2013-2016 for surgery residents (Sres), urology residents (Ures), pediatric surgery fellows (PSfel) and pediatric urology fellows (PUfel). Case log information for all levels of participation over all case categories that could potentially offer ERS volume were recorded. Volume was estimated using the mean number of included cases during residency and fellowship and the sum was used to estimate total training volume. Volume between groups was compared using the student's t test. RESULTS: Case logs were included for 4447 residents (4259 Sres, 840 Ures) and fellows (188 PSfel, 71 PUfel). Mean PU volume was 113.1, which was higher than the mean PS volume of 10.3 (p < 0.001). For PU, more ERS were performed during residency than fellowship (p < 0.001). For PS the opposite was true (p < 0.001). When examining fellow training only, PUfel performed more ERS than PSfel (11.7 vs. 7.0 p < 0.001). CONCLUSION: While previous publications note similar short-term outcomes for ERS for malignancy for PU and PS, ERS case volume during training is significantly different. Review of recent ACGME data indicate that PU have more overall experience with ERS, with most gained during residency. Additionally, PUfel performed significantly more ERS than PSfel. Further study into how these training differences affect long-term outcomes is necessary.


Assuntos
Bolsas de Estudo , Cirurgia Geral/educação , Internato e Residência , Nefrectomia/educação , Nefrectomia/estatística & dados numéricos , Pediatria/educação , Urologia/educação , Acreditação , Educação de Pós-Graduação em Medicina , Estados Unidos
2.
BMC Urol ; 19(1): 40, 2019 May 21.
Artigo em Inglês | MEDLINE | ID: mdl-31113422

RESUMO

BACKGROUND: Previously published results from our laboratory identified a mechano-gated two-pore domain potassium channel, TREK-1, as a main mechanosensor in the smooth muscle of the human urinary bladder. One of the limitations of in vitro experiments on isolated human detrusor included inability to evaluate in vivo effects of TREK-1 on voiding function, as the channel is also expressed in the nervous system, and may modulate micturition via neural pathways. Therefore, in the present study, we aimed to assess the role of TREK-1 channel in bladder function and voiding patterns in vivo by using TREK-1 knockout (KO) mice. METHODS: Adult C57BL/6 J wild-type (WT, N = 32) and TREK-1 KO (N = 33) mice were used in this study. The overall phenotype and bladder function were evaluated by gene and protein expression of TREK-1 channel, in vitro contractile experiments using detrusor strips in response to stretch and pharmacological stimuli, and cystometry in unanesthetized animals. RESULTS: TREK-1 KO animals had an elevated basal muscle tone and enhanced spontaneous activity in the detrusor without detectable changes in bladder morphology/histology. Stretch applied to isolated detrusor strips increased the amplitude of spontaneous contractions by 109% in the TREK-1 KO group in contrast to a 61% increase in WT mice (p ≤ 0.05 to respective baseline for each group). The detrusor strips from TREK-1 KO mice also generated more contractile force in response to electric field stimulation and high potassium concentration in comparison to WT group (p ≤ 0.05 for both tests). However, cystometric recordings from TREK-1 KO mice revealed a significant increase in the duration of the intermicturition interval, enhanced bladder capacity and increased number of non-voiding contractions in comparison to WT mice. CONCLUSIONS: Our results provide evidence that global down-regulation of TREK-1 channels has dual effects on detrusor contractility and micturition patterns in vivo. The observed differences are likely due to expression of TREK-1 channel not only in detrusor myocytes but also in afferent and efferent neural pathways involved in regulation of micturition which may underly the "mixed" voiding phenotype in TREK-1 KO mice.


Assuntos
Contração Muscular/fisiologia , Canais de Potássio de Domínios Poros em Tandem/deficiência , Bexiga Urinária/fisiologia , Micção/fisiologia , Animais , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout
3.
Am J Physiol Renal Physiol ; 313(2): F535-F546, 2017 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-28539337

RESUMO

Detrusor overactivity (DO) is the abnormal response of the urinary bladder to physiological stretch during the filling phase of the micturition cycle. The mechanisms of bladder smooth muscle compliance upon the wall stretch are poorly understood. We previously reported that the function of normal detrusor is regulated by TREK-1, a member of the mechanogated subfamily of two-pore-domain potassium (K2P) channels. In the present study, we aimed to identify the changes in expression and function of TREK-1 channels under pathological conditions associated with DO, evaluate the potential relationship between TREK-1 channels and cytoskeletal proteins in the human bladder, and test the possibility of modulation of TREK-1 channel expression by small RNAs. Expression of TREK-1 channels in DO specimens was 2.7-fold decreased compared with control bladders and was associated with a significant reduction of the recorded TREK-1 currents. Isolated DO muscle strips failed to relax when exposed to a TREK-1 channel opener. Immunocytochemical labeling revealed close association of TREK-1 channels with cell cytoskeletal proteins and caveolins, with caveolae microdomains being severely disrupted in DO specimens. Small activating RNA (saRNA) tested in vitro provided evidence that expression of TREK-1 protein could be partially upregulated. Our data confirmed a significant downregulation of TREK-1 expression in human DO specimens and provided evidence of close association between the channel, cell cytoskeleton, and caveolins. Upregulation of TREK-1 expression by saRNA could be a future step for the development of in vivo pharmacological and genetic approaches to treat DO in humans.


Assuntos
Canais de Potássio de Domínios Poros em Tandem/metabolismo , Bexiga Urinária Hiperativa/metabolismo , Caveolinas/metabolismo , Citoesqueleto/metabolismo , Regulação para Baixo , Humanos , Miócitos de Músculo Liso/metabolismo , RNA Interferente Pequeno
5.
J Biol Chem ; 287(35): 29968-78, 2012 Aug 24.
Artigo em Inglês | MEDLINE | ID: mdl-22761428

RESUMO

Emerging evidence suggests that the SAM pointed domain containing ETS transcription factor (SPDEF) plays a significant role in tumorigenesis in prostate, breast, colon, and ovarian cancer. However, there are no in vivo studies with respect to the role of SPDEF in tumor metastasis. The present study examined the effects of SPDEF on tumor cell metastasis using prostate tumor cells as a model. Utilizing two experimental metastasis models, we demonstrate that SPDEF inhibits cell migration and invasion in vitro and acts a tumor metastasis suppressor in vivo. Using stable expression of SPDEF in PC3-Luc cells and shRNA-mediated knockdown of SPDEF in LNCaP-Luc cells, we demonstrate for the first time that SPDEF diminished the ability of disseminated tumors cells to survive at secondary sites and establish micrometastases. These effects on tumor metastasis were not a result of the effect of SPDEF on cell growth as SPDEF expression had no effect on cell growth in vitro or subcutaneous tumor xenograft-growth in vivo. Transcriptional analysis of several genes associated with tumor metastasis, invasion, and the epithelial-mesenchymal transition demonstrated that SPDEF expression selectively down-regulated MMP9 and MMP13 in prostate cancer cells. Further analysis indicated that forced MMP9 or MMP13 expression rescued the invasive phenotype in SPDEF expressing PC3 cells in vitro, suggesting that the effects of SPDEF on tumor invasion are mediated, in part, through the suppression of MMP9 and MMP13 expression. These results demonstrate for the first time, in any system, that SPDEF functions as a tumor metastasis suppressor in vivo.


Assuntos
Movimento Celular , Neoplasias da Próstata/metabolismo , Proteínas Proto-Oncogênicas c-ets/metabolismo , Animais , Linhagem Celular Tumoral , Regulação para Baixo , Transição Epitelial-Mesenquimal/genética , Regulação Enzimológica da Expressão Gênica/genética , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Masculino , Metaloproteinase 13 da Matriz/biossíntese , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 9 da Matriz/biossíntese , Metaloproteinase 9 da Matriz/genética , Camundongos , Camundongos Nus , Invasividade Neoplásica , Metástase Neoplásica , Transplante de Neoplasias , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-ets/genética , Transcrição Gênica/genética , Transplante Heterólogo
7.
J Surg Res ; 172(1): 11-7, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-21777923

RESUMO

OBJECTIVES: In response to ACGME work-hour restrictions, residency programs that require continuous inpatient clinical care for educational objectives will be forced to increase the proportion of junior resident experience involved in shift work. Maintaining the balance of education over service at these levels will be a challenge, where a considerable amount of time must be spent gathering data for morning rounds and signing out patients at shift change. Patient safety is an issue with this new paradigm. We hypothesized that computerized sign-out would improve resident efficiency. MATERIALS AND METHODS: A multidisciplinary clinical team collaborated to design a computerized rounding and sign-out (CSO) program to automate collection of clinical information in addition to a brief narrative describing ongoing care issues. Residents returned a self-administered questionnaire before (n = 168) and after implementation (n = 83) examining: pre-rounding time, missed patients, handoff quality, and duty hours. RESULTS: Residents reported spending 11 fewer min/d pre-rounding (P = 0.006). After implementation, residents missed fewer patients on rounds (P = 0.01). A majority (70%) of responders stated that the new program helped them with duty hours. CONCLUSION: The current study demonstrates the reproducibility of the University of Washington model system for rounding and sign-out at an independent site, using basic infrastructure and leadership common to all residency programs. Developing a CSO was associated with a modest reduction in pre-rounding time and fewer patients missed on rounds. Although automating resident tasks may improve workflow in an increasingly complex hospital environment, structured handoff education and other institutional changes are necessary.


Assuntos
Simulação por Computador , Continuidade da Assistência ao Paciente/tendências , Internato e Residência , Administração dos Cuidados ao Paciente/tendências , Software/tendências , Eficiência , Humanos , Equipe de Assistência ao Paciente , Reprodutibilidade dos Testes , Segurança , Inquéritos e Questionários , Carga de Trabalho
8.
Surgery ; 172(5): 1407-1414, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36088172

RESUMO

BACKGROUND: Excess postoperative opioid prescribing increases the risk of opioid abuse, diversion, and addiction. Clinicians receive variable training for opioid prescribing, and despite the availability of guidelines, wide variations in prescribing practices persist. This quality improvement initiative aimed to assess and improve institutional adherence to published guidelines. METHODS: This study represented a quality improvement initiative at an academic medical center implemented over a 6-month period with data captured 1 year before and after implementation. The quality improvement initiative focused on prescribing education and monthly feedback reports for clinicians. All opioid-naïve, adult patients undergoing a reviewed procedure were included. Demographics, surgical details, hospital course, and opioid prescriptions were reviewed. Opioids prescribed on discharge were evaluated for concordance with recommendations based on published guidelines. Pre- and postimplementation cohorts were compared. RESULTS: There were 4,905 patients included: 2,343 preimplementation and 2,562 postimplementation. There were similar distributions in patient demographics between the 2 cohorts. Guideline-concordant discharge prescriptions improved from 50.3% to 72.2% after the quality improvement initiative was implemented (P < .001). Adjusted analysis controlling for sex, age, discharge clinician, length of stay, outpatient surgery, and procedure demonstrated a 190% increase in odds of receiving a guideline-concordant opioid prescription on discharge in the postimplementation cohort (adjusted odds ratio 2.90; 95% confidence interval = 2.55-3.30). CONCLUSION: This study represented a successful quality improvement initiative improving guideline-concordant opioid discharges and decreasing overprescribing. This study suggested published guidelines are insufficient without close attention to elements of effective change management including the critical importance of locally targeting educational efforts and suggested that real-time, data-driven feedback amplifies impact on prescribing behavior.


Assuntos
Analgésicos Opioides , Transtornos Relacionados ao Uso de Opioides , Adulto , Analgésicos Opioides/uso terapêutico , Fidelidade a Diretrizes , Humanos , Dor Pós-Operatória/tratamento farmacológico , Padrões de Prática Médica
9.
Am J Surg ; 223(1): 120-125, 2022 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-34407917

RESUMO

INTRODUCTION: Post-procedural debrief is recommended to improve patient safety. We examined operating room (OR) clinicians' perceptions of the impact of a multi-disciplinary debrief on OR culture. METHODS: A survey was administered to 182 OR clinicians at a major academic medical center. Attitudes toward the surgical debrief and its effect on patient safety and OR culture were evaluated. RESULTS: Majority of clinicians (58.2%) believed creating a culture of safety in the OR was a shared care team responsibility, however, surgical attendings and trainees were more likely to assign this responsibility to the surgical attending. Few circulating nurses and trainees felt comfortable initiating a surgical debrief. Overall clinicians agreed that a debrief would impact both patient safety outcomes and OR culture. CONCLUSIONS: Clinicians felt implementation of a surgical debrief would positively affect the OR culture of safety by improving interdisciplinary communication and influencing the power hierarchy that exists in many ORs.


Assuntos
Lista de Checagem/normas , Comunicação Interdisciplinar , Salas Cirúrgicas/organização & administração , Equipe de Assistência ao Paciente/organização & administração , Segurança do Paciente , Adulto , Feminino , Humanos , Masculino , Salas Cirúrgicas/normas , Cultura Organizacional , Equipe de Assistência ao Paciente/normas , Melhoria de Qualidade , Inquéritos e Questionários
10.
J Urol ; 186(6): 2426-33, 2011 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-22019171

RESUMO

PURPOSE: Histone deacetylase inhibitors represent promising cancer treatments since they offer improved access to target DNA/protein complexes by cytotoxic agents. We hypothesized that histone deacetylase inhibitors would be most effective when combined with DNA damaging agents such as mitomycin C. Valproic acid is a safe, affordable histone deacetylase inhibitor. We examined the effect of the combination of valproic acid and mitomycin C on human bladder cancer cells in vitro and compared this to the effect of valproic acid or mitomycin C alone on the cells. MATERIALS AND METHODS: We used HTB5 and HTB9 cells derived from low and high grade bladder tumors, respectively. HTB5 and HTB9 cells were grown in modified Eagle's and RPMI medium, respectively. Cell growth and proliferation were measured by standard methods. Apoptosis was evaluated microscopically after dual staining of cells with annexin V-fluorescein isothiocyanate/propidium iodide. The change in protein expression was analyzed by Western blot. RESULTS: Treatment of HTB5 and HTB9 bladder cancer cells for 24 to 72 hours with valproic acid and mitomycin C resulted in concentration and time dependent decreases in viability and proliferation. HTB9 cells showed marked sensitivity to mitomycin C with a 48-hour 50% median inhibitory concentration of 1 µg. Cells were less sensitive to valproic acid alone with a 48-hour 50% median inhibitory concentration of 2.5 mM. The chromatin structure relaxation induced by valproic acid pretreatment sensitized the bladder cancer cell lines, augmenting the cytotoxic action of mitomycin C. Valproic acid potentiated the induction of cell death by mitomycin C in each cell line in synergistic fashion. The effect of combining the 2 drugs was greater than the sum effect of each drug alone. CONCLUSIONS: Results indicate that the valproic acid and mitomycin C combination is effective, likely due to synergistic mechanisms. Animal model validation is needed but early results suggest promising intravesical treatments for superficial bladder cancer.


Assuntos
Antibióticos Antineoplásicos/uso terapêutico , Carcinoma de Células de Transição/tratamento farmacológico , Inibidores de Histona Desacetilases/uso terapêutico , Mitomicina/uso terapêutico , Neoplasias da Bexiga Urinária/tratamento farmacológico , Ácido Valproico/uso terapêutico , Sinergismo Farmacológico , Quimioterapia Combinada , Humanos , Células Tumorais Cultivadas
11.
Mol Cancer ; 9: 148, 2010 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-20550708

RESUMO

BACKGROUND: Prostate-derived Ets factor (PDEF) is expressed in tissues of high epithelial content including prostate, although its precise function has not been fully established. Conventional therapies produce a high rate of cure for patients with localized prostate cancer, but there is, at present, no effective treatment for intervention in metastatic prostate cancer. These facts underline the need to develop new approaches for early diagnosis of aggressive prostate cancer patients, and mechanism based anti-metastasis therapies that will improve the outlook for hormone-refractory prostate cancer. In this study we evaluated role of prostate-derived Ets factor (PDEF) in prostate cancer. RESULTS: We observed decreased PDEF expression in prostate cancer cell lines correlated with increased aggressive phenotype, and complete loss of PDEF protein in metastatic prostate cancer cell lines. Loss of PDEF expression was confirmed in high Gleason Grade prostate cancer samples by immuno-histochemical methods. Reintroduction of PDEF profoundly affected cell behavior leading to less invasive phenotypes in three dimensional cultures. In addition, PDEF expressing cells had altered cell morphology, decreased FAK phosphorylation and decreased colony formation, cell migration, and cellular invasiveness. In contrast PDEF knockdown resulted in increased migration and invasion as well as clonogenic activity. Our results also demonstrated that PDEF downregulated MMP9 promoter activity, suppressed MMP9 mRNA expression, and resulted in loss of MMP9 activity in prostate cancer cells. These results suggested that loss of PDEF might be associated with increased MMP9 expression and activity in aggressive prostate cancer. To confirm results we investigated MMP9 expression in clinical samples of prostate cancer. Results of these studies show increased MMP9 expression correlated with advanced Gleason grade. Taken together our results demonstrate decreased PDEF expression and increased MMP9 expression during the transition to aggressive prostate cancer. CONCLUSIONS: These studies demonstrate for the first time negative regulation of MMP9 expression by PDEF, and that PDEF expression was lost in aggressive prostate cancer and was inversely associated with MMP9 expression in clinical samples of prostate cancer. Based on these exciting results, we propose that loss of PDEF along with increased MMP9 expression should serve as novel markers for early detection of aggressive prostate cancer.


Assuntos
Metaloproteinase 9 da Matriz/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-ets/fisiologia , Sequência de Bases , Western Blotting , Linhagem Celular Tumoral , Primers do DNA , Humanos , Imuno-Histoquímica , Masculino , Fenótipo , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/metabolismo , Proteínas Proto-Oncogênicas c-ets/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa
12.
Mol Cancer Res ; 6(10): 1639-48, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18922979

RESUMO

Overexpression of focal adhesion kinase (FAK) has been well correlated with tumor development and/or the maintenance of tumor phenotype. In addition, inappropriate activation of the extracellular regulated kinase (ERK) signaling pathway is common to many human cancers. In the present study, we investigated the interplay between FAK and ERK in androgen-independent prostate cancer cells (PC3 and DU145 cells). We observed that suppression of FAK expression using small interfering RNA-mediated knockdown decreased the clonogenic activity, whereas overexpression of FAK increased it. We also observed that detachment of PC3 and DU145 cells from their substrate induced tyrosine phosphorylation of FAK. ERK knockdown diminished FAK protein levels and tyrosine phosphorylation of FAK as well as FAK promoter-reporter activity. We also tested the effect of MEK inhibitors and small interfering RNA-mediated knockdown of ERK1 and/or ERK2 on cell proliferation, invasiveness, and growth in soft agar of PC3 and DU145 cells. Inhibition of ERK signaling grossly impaired clonogenicity as well as invasion through Matrigel. However, inhibition of ERK signaling resulted in only a modest inhibition of 3H-thymidine incorporation and no effect on overall viability of the cells or increased sensitivity to anoikis. Taken together, these data show, for the first time, a requirement for FAK in aggressive phenotype of prostate cancer cells; reveal interdependence of FAK and ERK1/2 for clonogenic and invasive activity of androgen-independent prostate cancer cells; suggest a role for ERK regulation of FAK in substrate-dependent survival; and show for the first time, in any cell type, the regulation of FAK expression by ERK signaling pathway.


Assuntos
Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Neoplasias da Próstata/enzimologia , Neoplasias da Próstata/patologia , Androgênios/metabolismo , Anoikis , Linhagem Celular Tumoral , Proliferação de Células , Sobrevivência Celular , Ativação Enzimática , Humanos , Sistema de Sinalização das MAP Quinases , Masculino , Metaloproteinase 9 da Matriz/metabolismo , Proteína Quinase 1 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/antagonistas & inibidores , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , Invasividade Neoplásica , Fenótipo , Fosforilação , RNA Interferente Pequeno/metabolismo , Ensaio Tumoral de Célula-Tronco
13.
J Urol ; 179(5): 2057-63, 2008 May.
Artigo em Inglês | MEDLINE | ID: mdl-18355855

RESUMO

PURPOSE: Our understanding of physiological and pathophysiological events associated with inner medullary collecting duct epithelium is based on studies in cells isolated from mice and rats. We established primary cultures of hIMCD (human papillary collecting duct epithelial) cells. MATERIALS AND METHODS: Normal papillary tissues were dissected from the surgical waste of consenting patients undergoing renal surgery. Tissues were digested enzymatically. Cells were maintained in Dulbecco's modified Eagle's medium supplemented with glucose and antibiotics. Cultures were treated with ethylenediaminetetraacetic acid and epithelial select medium was also used to obtain a pure epithelial culture. RESULTS: The hIMCD cells grew in a monolayer. Cells showed the expression of epithelial specific markers, including cytokeratin, the tight junction marker zonula occludens 1 and the cytoskeletal protein vimentin. They lacked expression of factor VIII, which is a glycoprotein synthesized by endothelial cells. To our knowledge we also noted for the first time uroplakin expression in collecting duct epithelial cells. This expression was maintained in primary culture. The hIMCD cells in culture were highly resistant to hypertonic solutions and they responded to hypertonicity by cyclooxygenase-2 over expression. Moreover, these cells also survived prolonged periods of hypoxia. CONCLUSIONS: To our knowledge this is the first report of successful culture and characterization of primary cultures of collecting duct epithelial cells from human renal papillae. These cells will serve as essential tools in helping us fill the gaps in our understanding of the events associated with the physiology and pathophysiology of human renal inner medullary collecting duct epithelium.


Assuntos
Técnicas de Cultura de Células , Medula Renal/citologia , Túbulos Renais Coletores/citologia , Hipóxia Celular , Sobrevivência Celular , Meios de Cultura , Ciclo-Oxigenase 2/genética , Células Epiteliais/citologia , Expressão Gênica , Gliceraldeído 3-Fosfato Desidrogenase (NADP+)/genética , Humanos , Concentração Osmolar
14.
Curr Urol Rep ; 9(6): 500-5, 2008 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-18947516

RESUMO

Late-onset hypogonadism (LOH) and testosterone replacement therapy (TRT) are subjects of much recent research. Because aging men are at risk for benign prostatic hyperplasia (BPH) and prostate cancer, elucidating the relationship between testosterone and these diseases is crucial to ensure its safe administration. It is known that testosterone supplementation may worsen active prostate cancer and that its blockade or removal slows the disease's progression. However, recent studies have attempted to show that, in individuals in whom prostate cancer has been ruled out, TRT may simply restore serum testosterone levels to within normal limits without significant adverse affects on the prostate. Patients undergoing TRT should be monitored carefully for any evidence of prostatic disease.


Assuntos
Hipogonadismo/tratamento farmacológico , Próstata/fisiologia , Testosterona/uso terapêutico , Humanos , Masculino , Próstata/efeitos dos fármacos , Hiperplasia Prostática/etiologia , Neoplasias da Próstata/etiologia , Testosterona/farmacologia , Testosterona/fisiologia
15.
Front Syst Neurosci ; 12: 69, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30687029

RESUMO

Early activation of transcription factors is one of the epigenetic mechanisms contributing to the induction and maintenance of chronic pain states. Previous studies identified the changes in a number of nociception-related genes, such as calcitonin gene-related peptide (CGRP), substance P (SP), and brain-derived neurotropic factor (BDNF) in the pelvic organs after transient colonic inflammation. The gene and protein expression of these neuropeptides could be modulated by transcription factors Methyl-CpG-binding protein 2 (Mecp2) and cAMP response element-binding protein (CREB). In this study, we aimed to evaluate time-dependent changes in the expression levels of Mecp2 and CREB in the lumbosacral (LS) spinal cord and sensory ganglia after inflammation-induced pelvic pain in rat. Adult Sprague-Dawley rats were treated with 2,4,6-trinitrobenzenesulfonic acid (TNBS) to induce transient colonic inflammation. LS (L6-S2) spinal cord segments and respective dorsal root ganglias (DRGs) were isolated from control and experimental animals at 1, 2, 6, 24 h and 3 days post-TNBS treatment. Immunohistochemical (IHC) labeling and Western blotting experiments were performed to assess the expression of Mecp2, CREB and their phosphorylated forms. Total Mecp2 expression, but not phosphorylated p-Mecp2 (pS421Mecp2) expression was detected in the cells of the spinal dorsal horn under control conditions. Colonic inflammation triggered a significant decrease in the number of Mecp2-expressing neurons in parallel with elevated numbers of pS421Mecp2-expressing cells at 2 h and 6 h post-TNBS. The majority of Mecp2-positive cells (80 ± 6%) co-expressed CREB. TNBS treatment caused a transient up-regulation of CREB-expressing cells at 1 h post-TNBS only. The number of cells expressing phosphorylated CREB (pS133CREB) did not change at 1 h and 2 h post-TNBS, but was down-regulated by three folds at 6 h post-TNBS. Analysis of DRG sections revealed that the number of Mecp2-positive neurons was up-regulated by TNBS treatment, reaching three-fold increase at 2 h post-TNBS, and eight-fold increase at 6 h post-TNBS (p ≤ 0.05 to control). These data showed early changes in Mecp2 and CREB expression in the dorsal horn of the spinal cord and sensory ganglia after colonic inflammation, suggesting a possible contribution Mecp2 and CREB signaling in the development of visceral hyperalgesia and pelvic pain following peripheral inflammation.

16.
J Vis Exp ; (129)2017 11 02.
Artigo em Inglês | MEDLINE | ID: mdl-29155777

RESUMO

Transurethral instillation can be used to deliver different solutions with active ingredients (e.g., drugs, chemicals, bacteria, and viruses) locally into the urinary bladder to either induce animal models of bladder pathologies or evaluate the effectiveness of intravesical treatments. Most rodent models of lower urinary tract (LUT) pathologies are induced in female mice due to ease of intravesical instillation of the substances via the female urethra. However, due to anatomical differences between the female and male LUT, transurethral instillation in a male mouse has been deemed a very challenging procedure, and it has not been previously described. In this manuscript, we provide a detailed description of how to prepare polyethylene (PE) tubing for subsequent insertion into the urethra of a male mouse. In addition, we discuss the ideal types of PE tubing to be used depending on the desired site of inoculation. Furthermore, we describe point by point how to prepare an animal for a successful transurethral instillation to avoid injury to the urethra and ensure the delivery of the solution to the desired location. The procedure is started by retracting the prepuce and the glans to expose the opening of the urethral meatus. Next, the glans are grasped by blunt non-crushing forceps to stabilize the penis and the PE tubing. The PE tubing is first inserted into the urethral meatus parallel to the animal body, then its angle is adjusted by tilting the catheter to maneuver it to follow the natural curvature of the urethra. This technique can be used to induced murine models of bladder pathologies and/or evaluate the effectiveness of intravesical treatments in male mice.


Assuntos
Administração Intravesical , Uretra/cirurgia , Adulto , Animais , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL
17.
Urology ; 109: 223.e9-223.e16, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28827195

RESUMO

OBJECTIVE: To evaluate the differences in the composition and quantities of urine peptides in regular cannabis users and nonusers by liquid chromatography tandem mass spectrometry analysis. MATERIALS AND METHODS: Urine specimens from healthy control subjects and cannabis users were utilized to identify the differences in the number and quantity of urine proteins by liquid chromatography tandem mass spectrometry analysis. Significantly altered proteins were determined by a permutation testing statistical method. Heat map, dendrogram, pathway, and network analyses were performed to assess the degree of expression and the potential relationships between proteins in both groups. RESULTS: A total of 1337 proteins were detected in both groups with 19 proteins being significantly altered in cannabis users. Innate immunity and carbohydrate metabolic pathways were highly linked with upregulated proteins in the cannabis group. Additionally, 91 proteins were present and 46 proteins were absent only in cannabis users in comparison with the control cohort. Our results suggest that regular use of cannabis is associated with significant alterations in a number of urinary peptides, with a large number of proteins present or absent only in cannabis users. Pathway analyses demonstrated an increased immune response in cannabis users compared with controls. CONCLUSION: Our observations potentially indicate activation (or inhibition) of specific signaling pathways in the lower urinary tract during chronic exposure to exogenous cannabinoids. Our study provides initial proteomic knowledge for future investigations on the potential role of exocannabinoids in the development of intravesical therapies to treat lower urinary tract disorders.


Assuntos
Biomarcadores/urina , Canabinoides/farmacologia , Uso da Maconha/urina , Bexiga Urinária/efeitos dos fármacos , Bexiga Urinária/metabolismo , Cromatografia Líquida , Feminino , Humanos , Masculino , Uso da Maconha/metabolismo , Projetos Piloto , Proteômica
20.
Rev Urol ; 17(4): 211-9, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-26839518

RESUMO

Approximately 0.2% of Americans aged 20 to 39 years are childhood cancer survivors. Advances in cancer detection and therapy have greatly improved survival rates for young cancer patients; however, treatment of childhood cancers can adversely impact reproductive function. Many cancer patients report a strong desire to be informed of existing options for fertility preservation and future reproduction prior to initiation of gonadotoxic cancer therapies, including surgery, chemotherapy, and radiotherapy. This article discusses, in detail, the effects of cancer treatment on fertility in men and women, and outlines both current and experimental methods of fertility preservation among cancer patients.

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