RESUMO
STUDY OBJECTIVES: A growing body of literature suggests that deep brain stimulation (DBS) to treat motor symptoms of Parkinson's disease (PD) may also ameliorate certain sleep deficits. Many foundational studies have examined the impact of stimulation on sleep following several months of therapy, leaving an open question regarding the time course for improvement. It is unknown whether sleep improvement will immediately follow onset of therapy or accrete over a prolonged period of stimulation. The objective of our study was to address this knowledge gap by assessing the impact of DBS on sleep macro-architecture during the first nights of stimulation. METHODS: Polysomnograms were recorded for three consecutive nights in 14 patients with advanced PD (10 male, 4 female; age: 53-74 years), with intermittent, unilateral subthalamic nucleus DBS on the final night or two. Sleep scoring was determined manually by a consensus of four experts. Sleep macro-architecture was objectively quantified using the percentage, latency, and mean bout length of wake after sleep onset (WASO) and on each stage of sleep (REM and NREM stages N1, N2, N3). RESULTS: Sleep was found to be highly disrupted in all nights. Sleep architecture on nights without stimulation was consistent with prior results in treatment naive patients with PD. No statistically significant difference was observed due to stimulation. CONCLUSIONS: These objective measures suggest that one night of intermittent subthreshold stimulation appears insufficient to impact sleep macro-architecture. CLINICAL TRIAL REGISTRATION: Name: Adaptive Neurostimulation to Restore Sleep in Parkinson's Disease; URL: https://clinicaltrials.gov/ct2/show/NCT04620551; Identifier: NCT04620551.
RESUMO
OBJECTIVE: Sleep dysregulation in Parkinson's disease (PD) has been hypothesized to occur, in part, from dysfunction in the basal ganglia-cortical circuit. Assessment of this relationship requires accurate sleep stage determination, a known challenge in this clinical population. Our objective was to optimize the consensus on the sleep staging process and reduce interrater variability in a cohort of advanced PD subjects. METHODS: Fifteen PD subjects were enrolled from three sites in a clinical trial that involved recordings from subthalamic nucleus (STN) deep brain stimulation (DBS) leads (NCT04620551). Video polysomnography (vPSG) data for a total of 45 nights were analyzed. Four experienced scorers independently scored data on initial review. Epochs with less than 75% consensus were flagged for secondary review. In secondary review of discordant epochs, two of the original scorers re-assessed epochs, from which the final consensus stage was derived. RESULTS: Sleep stage classification agreement averaged 83.10% across all sleep stages on initial scoring (IS), and on secondary consensus scoring (CS) review, agreement reached 96.58%. Greatest disagreement was noted in determination of awake epochs (33.6% of discordant epochs) and non-rapid-eye-movement stage 2 (N2) epochs (31.8% of discordant epochs). Scoring discrepancy was resolved with direct measurement of cortical frequency and amplitudes, physiologic context of the epoch, and video review. CONCLUSION: Our method of multi-level initial and then secondary consensus review scoring resulted in consensus scoring agreement superior to conventional standards. This work features a custom-engineered vPSG software and review platform for integration of consensus sleep stage scoring in a multi-site clinical trial.
Assuntos
Doença de Parkinson , Humanos , Consenso , Variações Dependentes do Observador , Doença de Parkinson/complicações , Reprodutibilidade dos Testes , Sono , Fases do Sono/fisiologiaRESUMO
Activation of the NF-κB pathway is causally linked to initiation and progression of diverse cancers. Therefore, IKKß, the key regulatory kinase of the canonical NF-κB pathway, should be a logical target for cancer treatment. However, existing IKKß inhibitors are known to induce paradoxical immune activation, which limits their clinical usefulness. Recently, we identified a quinoxaline urea analog 13-197 as a novel IKKß inhibitor that delays tumor growth without significant adverse effects in xenograft tumor models. In the present study, we found that 13-197 had little effect on LPS-induced NF-κB target gene induction by primary mouse macrophages while maintaining considerable anti-proliferative activities. These characteristics may explain absence of inflammatory side effects in animals treated with 13-197. Our data also demonstrate that the inflammation and proliferation-related functions of IKKß can be uncoupled, and highlight the utility of 13-197 to dissect these downstream pathways.
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Quinase I-kappa B/antagonistas & inibidores , Macrófagos/efeitos dos fármacos , Compostos de Fenilureia/farmacologia , Quinoxalinas/farmacologia , Animais , Western Blotting , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Citocinas/genética , Citocinas/imunologia , Citocinas/metabolismo , Citometria de Fluxo , Expressão Gênica/efeitos dos fármacos , Quinase I-kappa B/imunologia , Quinase I-kappa B/metabolismo , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Lipopolissacarídeos/farmacologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Fosforilação/efeitos dos fármacos , Fosforilação/imunologiaRESUMO
Based on gene expression patterns, breast cancers can be divided into subtypes that closely resemble various developmental stages of normal mammary epithelial cells (MECs). Thus, understanding molecular mechanisms of MEC development is expected to provide critical insights into initiation and progression of breast cancer. Epidermal growth factor receptor (EGFR) and its ligands play essential roles in normal and pathological mammary gland. Signals through EGFR is required for normal mammary gland development. Ligands for EGFR are over-expressed in a significant proportion of breast cancers, and elevated expression of EGFR is associated with poorer clinical outcome. In the present study, we examined the effect of signals through EGFR on MEC differentiation using the human telomerase reverse transcriptase (hTERT)-immortalized human stem/progenitor MECs which express cytokeratin 5 but lack cytokeratin 19 (K5(+)K19(-) hMECs). As reported previously, these cells can be induced to differentiate into luminal and myoepithelial cells under appropriate culture conditions. K5(+)K19(-) hMECs acquired distinct cell fates in response to EGFR ligands epidermal growth factor (EGF), amphiregulin (AREG) and transforming growth factor alpha (TGFα) in differentiation-promoting MEGM medium. Specifically, presence of EGF during in vitro differentiation supported development into both luminal and myoepithelial lineages, whereas cells differentiated only towards luminal lineage when EGF was replaced with AREG. In contrast, substitution with TGFα led to differentiation only into myoepithelial lineage. Chemical inhibition of the MEK-Erk pathway, but not the phosphatidylinositol 3-kinase (PI3K)-AKT pathway, interfered with K5(+)K19(-) hMEC differentiation. The present data validate the utility of the K5(+)K19(-) hMEC cells for modeling key features of human MEC differentiation. This system should be useful in studying molecular/biochemical mechanisms of human MEC differentiation.
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Diferenciação Celular/fisiologia , Receptores ErbB/metabolismo , Glândulas Mamárias Humanas/citologia , Anfirregulina , Diferenciação Celular/genética , Linhagem Celular , Família de Proteínas EGF , Fator de Crescimento Epidérmico/farmacologia , Células Epiteliais , Receptores ErbB/agonistas , Citometria de Fluxo , Imunofluorescência , Glicoproteínas/farmacologia , Humanos , Immunoblotting , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Microscopia Confocal , Fator de Crescimento Transformador alfa/farmacologiaRESUMO
Luteolysis is characterized by angioregression, luteal cell apoptosis, and remodeling of the extracellular matrix characterized by deposition of collagen 1. Transforming growth factor beta 1 (TGFB1) is a potent mediator of wound healing and fibrotic processes through stimulation of the synthesis of extracellular matrix components. We hypothesized that TGFB1 stimulates profibrotic activities of luteal fibroblasts. We examined the actions of TGFB1 on luteal fibroblast proliferation, extracellular matrix production, floating gel contraction, and chemotaxis. Fibroblasts were isolated from the bovine corpus luteum. Western blot analysis showed that luteal fibroblasts expressed collagen 1 and prolyl 4-hydroxylase but did not express markers of endothelial or steroidogenic cells. Treatment of fibroblasts with TGFB1 stimulated the phosphorylation of SMAD2 and SMAD3. [(3)H]thymidine incorporation studies showed that TGFB1 caused concentration-dependent reductions in DNA synthesis in luteal fibroblasts and significantly (P < 0.05) reduced the proliferative effect of FGF2 and fetal calf serum. However, TGFB1 did not reduce the viability of luteal fibroblasts. Treatment of luteal fibroblasts with TGFB1 induced the expression of laminin, collagen 1, and matrix metalloproteinase 1 as determined by Western blot analysis and gelatin zymography of conditioned medium. TGFB1 increased the chemotaxis of luteal fibroblasts toward fibronectin in a transwell system. Furthermore, TGFB1 increased the fibroblast-mediated contraction of floating bovine collagen 1 gels. These results suggest that TGFB1 contributes to the structural regression of the corpus luteum by stimulating luteal fibroblasts to remodel and contract the extracellular matrix.
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Bovinos , Corpo Lúteo/citologia , Fibroblastos/efeitos dos fármacos , Fibroblastos/fisiologia , Fator de Crescimento Transformador beta1/farmacologia , Animais , Proliferação de Células , Sobrevivência Celular/efeitos dos fármacos , Quimiotaxia/efeitos dos fármacos , Colágeno/análise , DNA/biossíntese , Proteínas da Matriz Extracelular/biossíntese , Feminino , Fibroblastos/química , Fibrose , Laminina/análise , Metaloproteinases da Matriz/análise , Fosforilação/efeitos dos fármacos , Proteína Smad2/metabolismo , Proteína Smad3/metabolismoRESUMO
We assessed neonatal diethylstilbestrol (DES)-induced disruption at various endocrine levels in the hamster. In particular, we used organ transplantation into the hamster cheek pouch to determine whether abnormalities observed in the post-pubertal ovary are due to: (a) a direct (early) mechanism or (b) an indirect (late) mechanism that involves altered development and function of the hypothalamus and/or pituitary. Of the various disruption endpoints and attributes assessed: (1) some were consistent with the direct mechanism (altered uterine and cervical dimensions/organization, ovarian polyovular follicles, vaginal hypospadius, endometrial hyperplasia/dysplasia); (2) some were consistent with the indirect mechanism (ovarian/oviductal salpingitis, cystic ovarian follicles); (3) some were consistent with a combination of the direct and indirect mechanisms (altered endocrine status); and (4) the mechanism(s) for one (lack of corpora lutea) was uncertain. This study also generated some surprising observations regarding vaginal estrous assessments as a means to monitor periodicity of ovarian function in the hamster.
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Animais Recém-Nascidos , Dietilestilbestrol/toxicidade , Estrogênios não Esteroides/toxicidade , Genitália Feminina/efeitos dos fármacos , Animais , Colo do Útero/anatomia & histologia , Cricetinae , Ciclo Estral , Tubas Uterinas/anatomia & histologia , Tubas Uterinas/efeitos dos fármacos , Tubas Uterinas/fisiologia , Feminino , Genitália Feminina/anatomia & histologia , Genitália Feminina/fisiologia , Hormônios/sangue , Hipotálamo/fisiologia , Mesocricetus , Ovariectomia , Ovário/anatomia & histologia , Ovário/fisiologia , Ovário/transplante , Hipófise/fisiologia , Gravidez , Maturidade Sexual , Útero/anatomia & histologia , Vagina/fisiologiaRESUMO
Cyclical formation and regression of the ovarian corpus luteum is required for reproduction. During luteal regression, the microvasculature of the corpus luteum is extensively disrupted. Prostaglandin F2α, a primary signal for luteal regression, induces the expression of transforming growth factor ß1 (TGFB1) in the corpus luteum. This study determined the actions of TGFB1 on microvascular endothelial cells isolated from the bovine corpus luteum (CLENDO cells). We hypothesized that TGFB1 participates in the disruption of the microvasculature during luteal regression. TGFB1 activated the canonical SMAD signaling pathway in CLENDO cells. TGFB1 (1 ng/ml) significantly reduced both basal and fetal-calf-serum-stimulated DNA synthesis, without reducing cell viability. TGFB1 also significantly reduced CLENDO cell transwell migration and disrupted the formation of capillary-like structures when CLENDO cells were plated on Matrigel. By contrast, CLENDO cells plated on fibrillar collagen I gels did not form capillary-like structures and TGFB1 induced cell death. Additionally, TGFB1 caused loss of VE-cadherin from cellular junctions and loss of cell-cell contacts, and increased the permeability of confluent CLENDO cell monolayers. These studies demonstrate that TGFB1 acts directly on CLENDO cells to limit endothelial cell function and suggest that TGFB1 might act in the disassembly of capillaries observed during luteal regression.
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Corpo Lúteo/irrigação sanguínea , Células Endoteliais/metabolismo , Fator de Crescimento Transformador beta1/metabolismo , Animais , Bovinos , Movimento Celular/fisiologia , Células Endoteliais/citologia , Feminino , Humanos , Luteólise/metabolismo , Fosforilação , Transdução de Sinais , Fator de Crescimento Transformador beta1/genéticaRESUMO
BACKGROUND: Immunosuppression medications contribute to posttransplant diabetes mellitus in patients and can cause insulin resistance in male rats. Tacrolimus (TAC)-sirolimus (SIR) immunosuppression is also associated with appearance of ovarian cysts in transplant patients. Because insulin resistance is observed in patients with polycystic ovary syndrome, we hypothesized that TAC or SIR may induce reproductive abnormalities. METHODS: We monitored estrus cycles of adult female rats treated daily with TAC, SIR, and combination of TAC-SIR, or diluent (control) for 4 weeks. Animals were then challenged with oral glucose to determine their glucose and insulin responses, killed, and their blood and tissues, including ovaries and uteri harvested. RESULTS: TAC and TAC-SIR treatments increased mean random glucose concentrations (P<0.05). TAC, SIR, and TAC-SIR treatments also increased the glucose response to oral glucose challenge (P<0.05). The insulin response to glucose was significantly higher in rats treated with SIR compared with TAC (P<0.05). TAC, SIR and TAC-SIR treatments reduced number of estrus cycles (P<0.05). The ovaries were smaller after SIR and TAC-SIR treatment compared with controls. The TAC and TAC-SIR treatment groups had fewer preovulatory follicles. Corpora lutea were present in all groups. Ovarian aromatase expression was reduced in the SIR and TAC-SIR treatment groups. A significant (P<0.05) reduction in uterine size was observed in all treatment groups when compared with controls. CONCLUSION: In a model of immunosuppressant-induced hyperglycemia, both TAC and SIR induced reproductive abnormalities in adult female rats, likely through different mechanisms.
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Sirolimo/efeitos adversos , Tacrolimo/efeitos adversos , Animais , Aromatase/biossíntese , Glicemia/metabolismo , Estro , Feminino , Regulação Enzimológica da Expressão Gênica , Glucose/uso terapêutico , Hiperglicemia/induzido quimicamente , Imunossupressores/efeitos adversos , Resistência à Insulina , Ovário/efeitos dos fármacos , Fenótipo , Síndrome do Ovário Policístico/induzido quimicamente , Ratos , Ratos Sprague-Dawley , Útero/efeitos dos fármacosRESUMO
Progesterone secretion by the steroidogenic cells of the corpus luteum (CL) is essential for reproduction. Progesterone synthesis is under the control of LH, but the exact mechanism of this regulation is unknown. It is established that LH stimulates the LH receptor/choriogonadotropin receptor, a G-protein coupled receptor, to increase cAMP and activate cAMP-dependent protein kinase A (PKA). In the present study, we tested the hypothesis that cAMP/PKA-dependent regulation of the Wnt pathway components glycogen synthase kinase (GSK)-3beta and beta-catenin contributes to LH-dependent steroidogenesis in luteal cells. We observed that LH via a cAMP/PKA-dependent mechanism stimulated the phosphorylation of GSK3beta at N-terminal Ser9 causing its inactivation and resulted in the accumulation of beta-catenin. Overexpression of N-terminal truncated beta-catenin (Delta90 beta-catenin), which lacks the phosphorylation sites responsible for its destruction, significantly augmented LH-stimulated progesterone secretion. In contrast, overexpression of a constitutively active mutant of GSK3beta (GSK-S9A) reduced beta-catenin levels and inhibited LH-stimulated steroidogenesis. Chromatin immunoprecipitation assays demonstrated the association of beta-catenin with the proximal promoter of the StAR gene, a gene that expresses the steroidogenic acute regulatory protein, which is a cholesterol transport protein that controls a rate-limiting step in steroidogenesis. Collectively these data suggest that cAMP/PKA regulation of GSK3beta/beta-catenin signaling may contribute to the acute increase in progesterone production in response to LH.