RESUMO
Clostridioides difficile infection (CDI) causes nosocomial/antibiotic-associated gastrointestinal diseases with dramatically increasing global incidence and mortality rates. The main C. difficile virulence factors, toxins A and B (TcdA/TcdB), cause cytopathic/cytotoxic effects and inflammation. We demonstrated that TcdB induces caspase-dependent, mitochondria-independent enteric glial cell (EGC) apoptosis that is enhanced by the pro-inflammatory cytokines TNF-α and IFN-γ (CKs) by increasing caspase-3/7/9 and PARP activation. Because this cytotoxic synergism is important for CDI pathogenesis, we investigated the apoptotic pathways involved in TcdB- and TcdB + CK-induced apoptosis indepth. EGCs were pre-treated with the inhibitors BAF or Q-VD-OPh (pan-caspase), Z-DEVD-fmk (caspase-3/7), Z-IETD-fmk (caspase-8), PD150606 (calpains), and CA-074Me (cathepsin B) 1 h before TcdB exposure, while CKs were given 1.5 h after TcdB exposure, and assays were performed at 24 h. TcdB and TcdB + CKs induced apoptosis through three signalling pathways activated by calpains, caspases and cathepsins, which all are involved both in induction and execution apoptotic signalling under both conditions but to different degrees in TcdB and TcdB + CKs especially as regards to signal transduction mediated by these proteases towards downstream effects (apoptosis). Calpain activation by Ca2+ influx is the first pro-apoptotic event in TcdB- and TcdB + CK-induced EGC apoptosis and causes caspase-3, caspase-7 and PARP activation. PARP is also directly activated by calpains which are responsible of about 75% of apoptosis in TcdB and 62% in TcdB + CK which is both effector caspase-dependent and -independent. Initiator caspase-8 activation mediated by TcdB contributes to caspase-3/caspase-7 and PARP activation and is responsible of about 28% of apoptosis in both conditions. Caspase-3/caspase-7 activation is weakly responsible of apoptosis, indeed we found that it mediates 27% of apoptosis only in TcdB. Cathepsin B contributes to triggering pro-apoptotic signal and is responsible in both conditions of about 35% of apoptosis by a caspase-independent manner, and seems to regulate the caspase-3 and caspase-7 cleaved fragment levels, highlighting the complex interaction between these cysteine protease families activated during TcdB-induced apoptosis. Further a relevant difference between TcdB- and TcdB + CK-induced apoptosis is that TcdB-induced apoptosis increased slowly reaching at 72 h the value of 18.7%, while TcdB + CK-induced apoptosis increased strongly reaching at 72 h the value of 60.6%. Apoptotic signalling activation by TcdB + CKs is enriched by TNF-α-induced NF-κB signalling, inhibition of JNK activation and activation of AKT. In conclusion, the ability of C. difficile to activate three apoptotic pathways represents an important strategy to overcome resistance against its cytotoxic activity.
Assuntos
Toxinas Bacterianas , Clostridioides difficile , Infecções por Clostridium , Apoptose/fisiologia , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/toxicidade , Calpaína/metabolismo , Caspase 3/metabolismo , Caspase 7/metabolismo , Caspase 7/farmacologia , Caspases/metabolismo , Catepsina B/metabolismo , Citocinas/metabolismo , Humanos , Neuroglia/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo , Fator de Necrose Tumoral alfa/farmacologiaRESUMO
Cytokinesis is monitored by a molecular machinery that promotes the degradation of the intercellular bridge, a transient protein structure connecting the two daughter cells. Here, we found that CSA and CSB, primarily defined as DNA repair factors, are located at the midbody, a transient structure in the middle of the intercellular bridge, where they recruit CUL4 and MDM2 ubiquitin ligases and the proteasome. As a part of this molecular machinery, CSA and CSB contribute to the ubiquitination and the degradation of proteins such as PRC1, the Protein Regulator of Cytokinesis, to ensure the correct separation of the two daughter cells. Defects in CSA or CSB result in perturbation of the abscission leading to the formation of long intercellular bridges and multinucleated cells, which might explain part of the Cockayne syndrome phenotypes. Our results enlighten the role played by CSA and CSB as part of a ubiquitin/proteasome degradation process involved in transcription, DNA repair, and cell division.
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Divisão Celular , DNA Helicases/metabolismo , Enzimas Reparadoras do DNA/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Fatores de Transcrição/metabolismo , Ubiquitina/metabolismo , DNA Helicases/genética , Enzimas Reparadoras do DNA/genética , Imunofluorescência , Humanos , Mitose , Proteínas de Ligação a Poli-ADP-Ribose/genética , Ligação Proteica , Transporte Proteico , Proteólise , Fuso Acromático , Fatores de Transcrição/genética , UbiquitinaçãoRESUMO
We investigated the association between circulating microRNAs (miRNAs) potentially involved in the lung inflammatory process and fibrosis development among COVID-19-related acute respiratory distress syndrome (ARDS) survivors. At 4 ± 2 months from clinical recovery, COVID-19-related ARDS survivors matched for age, sex, and clinical characteristics underwent chest high-resolution computerized tomography (HRCT) and were selected based on imaging pattern evolution into fully recovered (N = normal), pulmonary opacities (PO) and fibrosis-like lesions (FL). Based on the previous literature, we performed plasma miRNA profiling of exosomal miRNAs belonging to the NLRP3-inflammasome platform with validated (miR-17-5p, miR-223-3p) and putative targets (miR-146a-5p), miRNAs involved in the post-transcriptional regulation of acute phase cytokines (miR128-3p, miR3168, miR125b-2-3p, miR106a-5p), miRNAs belonging to the NLRP4-inflammasome platform (miR-141-3p) and miRNAs related to post-transcriptional regulation of the fibrosis process (miR-21-5p). miR-17-5p, miR-223-3p, and miR-146a-5p were significantly down-regulated in patients with FL when compared to patients with PO. miR-146a-5p was also down-regulated in patients with FL than in N. The expression of the remaining miRNAs did not differ by group. In patients with long-term pulmonary radiological sequelae following COVID-19-related ARDS, a down-regulation of miR-17-5p, miR-146a-3p, and miR-223-3p correlated to fibrosis development in patients showing persistent hyper-reactivity to inflammatory stimulation. Our results support the hypothesis that NLRP3-Inflammasome could be implicated in the process of fibrotic evolution of COVID-19-associated ARDS.
Assuntos
COVID-19 , MicroRNAs , Síndrome do Desconforto Respiratório , Humanos , Inflamassomos , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , COVID-19/diagnóstico por imagem , COVID-19/genética , MicroRNAs/genética , Síndrome do Desconforto Respiratório/diagnóstico por imagem , Síndrome do Desconforto Respiratório/genética , Progressão da Doença , SobreviventesRESUMO
INTRODUCTION AND HYPOTHESIS: The aims of this study were to evaluate by transperineal ultrasound if there were ultrasound-detectable changes over time in the dynamic behavior of the sling in patients who underwent transobturator tape (TOT), and to evaluate if dynamic translabial ultrasonography recognized factors that may be associated with failed surgery. METHODS: This was a single-center prospective study. We included women who underwent "out-in" TOT for stress urinary incontinence (SUI). A dynamic translabial ultrasound was performed 6 months post-surgery and again at the last visit. The objective cure for SUI was defined as the absence of urine leakage during the stress test. We evaluated the bladder neck mobility at rest and during Valsalva; the position of the mesh along the urethra; the concordance of urethral movement with the sling during Valsalva; the symmetry of the lateral arms of the sling during straining; and the presence or absence of bladder neck funneling. RESULTS: From December 2012 to February 2016, 80 consecutive patients were included. Six months after surgery, incontinent women compared with continent women had the sling in a proximal or distal position, that moved discordantly with the urethra (p < 0.0001), with asymmetry arm and bladder neck funneling (p < 0.0001). Continent patients had a significant improvement of urethrocele grade both at rest (p = 0.036) and during Valsalva (p = 0.045). CONCLUSIONS: Technical and positioning errors can lead to the failure of anti-incontinence surgical treatment. Translabial ultrasound allows the correct positioning of the sling to be evaluated and any errors that need to be analyzed in order to then solve the failure.
Assuntos
Slings Suburetrais , Incontinência Urinária por Estresse , Feminino , Humanos , Masculino , Estudos Prospectivos , Ultrassonografia/métodos , Uretra/diagnóstico por imagem , Incontinência Urinária por Estresse/complicações , Incontinência Urinária por Estresse/diagnóstico por imagem , Incontinência Urinária por Estresse/cirurgiaRESUMO
Varicocele (VC) is the most common abnormality identified in men evaluated for hypofertility. Increased levels of reactive oxygen species (ROS) and reduced antioxidants concentrations are key contributors in varicocele-mediated hypofertility. Moreover, inflammation and alterations in testicular immunity negatively impact male fertility. In particular, NLRP3 inflammasome activation was hypothesized to lead to seminal inflammation, in which the levels of specific cytokines, such as IL-1ß and IL-18, are overexpressed. In this review, we described the role played by oxidative stress (OS), inflammation, and NLRP3 inflammasome activation in VC disease. The consequences of ROS overproduction in testis, including inflammation, lipid peroxidation, mitochondrial dysfunction, chromatin damage, and sperm DNA fragmentation, leading to abnormal testicular function and failed spermatogenesis, were highlighted. Finally, we described some therapeutic antioxidant strategies, with recognized beneficial effects in counteracting OS and inflammation in testes, as possible therapeutic drugs against varicocele-mediated hypofertility.
Assuntos
Varicocele , Antioxidantes/farmacologia , Humanos , Inflamassomos/metabolismo , Inflamação , Masculino , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Estresse Oxidativo , Espécies Reativas de Oxigênio/farmacologia , Varicocele/tratamento farmacológicoRESUMO
INTRODUCTION: The aim of this study was to assess whether antibiotic prophylaxis or therapy is sufficient for laparoscopic or vaginal prolapse surgery with mesh. METHODS: This is a single-center prospective study. The study was divided into 3 groups. Protocol A: metronidazole (15 mg/kg) and piperacillin-tazobactam (2 g) 1 h before surgery and, for postoperative treatment, gentamycin (160 mg) 1 h before surgery in a single dose. Metronidazole and piperacillin-tazobactam were administered until hospital discharge. Protocol B: gentamycin and piperacillin-tazobactam in the same manner as group A. Protocol C: clindamycin (600 mg) and gentamicin (160 mg) 1 h before surgery in a single dose. RESULTS: We included 87 consecutive patients who underwent prolapse surgery involving mesh prostheses: 57 by the laparoscopic approach and 30 by the vaginal route. Of these, 30 patients were included in protocol A, 30 in protocol B, and 27 in protocol C. There were no statistically significant differences among the 3 protocols regarding any postoperative complications, except for urinary tract infections that were more in the vaginal approach than in the laparoscopic route, in protocol A (p = 0.002). CONCLUSIONS: One-shot prophylaxis can be successfully used in prolapse surgery regardless of the surgical approach.
Assuntos
Antibacterianos/administração & dosagem , Antibioticoprofilaxia , Gentamicinas/administração & dosagem , Procedimentos Cirúrgicos em Ginecologia , Laparoscopia , Metronidazol/administração & dosagem , Prolapso de Órgão Pélvico/cirurgia , Combinação Piperacilina e Tazobactam/administração & dosagem , Infecção da Ferida Cirúrgica/prevenção & controle , Infecções Urinárias/prevenção & controle , Idoso , Antibacterianos/efeitos adversos , Antibioticoprofilaxia/efeitos adversos , Esquema de Medicação , Feminino , Gentamicinas/efeitos adversos , Procedimentos Cirúrgicos em Ginecologia/efeitos adversos , Procedimentos Cirúrgicos em Ginecologia/instrumentação , Humanos , Laparoscopia/efeitos adversos , Laparoscopia/instrumentação , Metronidazol/efeitos adversos , Pessoa de Meia-Idade , Combinação Piperacilina e Tazobactam/efeitos adversos , Estudos Prospectivos , Telas Cirúrgicas , Infecção da Ferida Cirúrgica/microbiologia , Fatores de Tempo , Resultado do Tratamento , Infecções Urinárias/microbiologiaRESUMO
Neuroblastoma, the most common extra-cranial solid tumor of early childhood, is one of the major therapeutic challenges in child oncology: it is highly heterogenic at a genetic, biological, and clinical level. The high-risk cases have one of the least favorable outcomes amongst pediatric tumors, and the mortality rate is still high, regardless of the use of intensive multimodality therapies. Here, we observed that neuroblastoma cells display an increased expression of Cockayne Syndrome group B (CSB), a pleiotropic protein involved in multiple functions such as DNA repair, transcription, mitochondrial homeostasis, and cell division, and were recently found to confer cell robustness when they are up-regulated. In this study, we demonstrated that RNAi-mediated suppression of CSB drastically impairs tumorigenicity of neuroblastoma cells by hampering their proliferative, clonogenic, and invasive capabilities. In particular, we observed that CSB ablation induces cytokinesis failure, leading to caspases 9 and 3 activation and, subsequently, to massive apoptotic cell death. Worthy of note, a new frontier in cancer treatment, already proved to be successful, is cytokinesis-failure-induced cell death. In this context, CSB ablation seems to be a new and promising anticancer strategy for neuroblastoma therapy.
Assuntos
Citocinese/fisiologia , DNA Helicases/fisiologia , Enzimas Reparadoras do DNA/fisiologia , Neuroblastoma/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose/fisiologia , Interferência de RNA , Apoptose , Caspase 3/metabolismo , Caspase 9/metabolismo , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Centrossomo , DNA Helicases/genética , DNA Helicases/metabolismo , Reparo do DNA , Enzimas Reparadoras do DNA/genética , Humanos , Proteínas de Ligação a Poli-ADP-Ribose/genética , Fuso AcromáticoRESUMO
HOPS is a ubiquitin-like protein implicated in many aspects of cellular function including the regulation of mitotic activity, proliferation, and cellular stress responses. In this study, we focused on the complex relationship between HOPS and the tumor suppressor p53, investigating both transcriptional and non-transcriptional p53 responses. Here, we demonstrated that Hops heterozygous mice and mouse embryonic fibroblasts exhibit an impaired DNA-damage response to etoposide-induced double-strand breaks when compared to wild-type genes. Specifically, alterations in HOPS levels caused significant defects in the induction of apoptosis, including a reduction in p53 protein level and percentage of apoptotic cells. We also analyzed the effect of reduced HOPS levels on the DNA-damage response by examining the transcript profiles of p53-dependent genes, showing a suggestive deregulation of the mRNA levels for a number of p53-dependent genes. Taken together, these results show an interesting haploinsufficiency effect mediated by Hops monoallelic deletion, which appears to be enough to destabilize the p53 protein and its functions. Finally, these data indicate a novel role for Hops as a tumor-suppressor gene in DNA damage repair in mammalian cells.
Assuntos
Apoptose , Reparo do DNA , Haploinsuficiência , Peptídeos e Proteínas de Sinalização Intracelular/fisiologia , Proteínas de Membrana/fisiologia , Proteína Supressora de Tumor p53/metabolismo , Animais , Dano ao DNA , Feminino , Heterozigoto , Masculino , CamundongosRESUMO
The ability of spermatozoa to swim towards an oocyte and fertilize it depends on precise K+ permeability changes. Kir5.1 is an inwardly-rectifying potassium (Kir) channel with high sensitivity to intracellular H+ (pHi) and extracellular K+ concentration [K+]o, and hence provides a link between pHi and [K+]o changes and membrane potential. The intrinsic pHi sensitivity of Kir5.1 suggests a possible role for this channel in the pHi-dependent processes that take place during fertilization. However, despite the localization of Kir5.1 in murine spermatozoa, and its increased expression with age and sexual maturity, the role of the channel in sperm morphology, maturity, motility, and fertility is unknown. Here, we confirmed the presence of Kir5.1 in spermatozoa and showed strong expression of Kir4.1 channels in smooth muscle and epithelial cells lining the epididymal ducts. In contrast, Kir4.2 expression was not detected in testes. To examine the possible role of Kir5.1 in sperm physiology, we bred mice with a deletion of the Kcnj16 (Kir5.1) gene and observed that 20% of Kir5.1 knock-out male mice were infertile. Furthermore, 50% of knock-out mice older than 3 months were unable to breed. By contrast, 100% of wild-type (WT) mice were fertile. The genetic inactivation of Kcnj16 also resulted in smaller testes and a greater percentage of sperm with folded flagellum compared to WT littermates. Nevertheless, the abnormal sperm from mutant animals displayed increased progressive motility. Thus, ablation of the Kcnj16 gene identifies Kir5.1 channel as an important element contributing to testis development, sperm flagellar morphology, motility, and fertility. These findings are potentially relevant to the understanding of the complex pHi- and [K+]o-dependent interplay between different sperm ion channels, and provide insight into their role in fertilization and infertility.
Assuntos
Infertilidade Masculina/genética , Canais de Potássio Corretores do Fluxo de Internalização/genética , Espermatozoides/metabolismo , Animais , Fertilidade/genética , Regulação da Expressão Gênica no Desenvolvimento/genética , Infertilidade Masculina/patologia , Masculino , Potenciais da Membrana/genética , Camundongos , Camundongos Knockout , Músculo Liso/metabolismo , Oócitos/crescimento & desenvolvimento , Potássio/metabolismo , Motilidade dos Espermatozoides/genética , Espermatozoides/crescimento & desenvolvimento , Testículo/crescimento & desenvolvimento , Testículo/metabolismo , Canal Kir5.1RESUMO
The innate immune system represents the host's first-line defense against pathogens, dead cells or environmental factors. One of the most important inflammatory pathways is represented by the activation of the NOD-like receptor (NLR) protein family. Some NLRs induce the assembly of large caspase-1-activating complexes called inflammasomes. Different types of inflammasomes have been identified that can respond to distinct bacterial, viral or fungal infections; sterile cell damage or other stressors, such as metabolic imbalances. Epigenetic regulation has been recently suggested to provide a complementary mechanism to control inflammasome activity. This regulation can be exerted through at least three main mechanisms, including CpG DNA methylation, histones post-translational modifications and noncoding RNA expression. The repression or promotion of expression of different inflammasomes (NLRP1, NLRP2, NLRP3, NLRP4, NLRP6, NLRP7, NLRP12 and AIM2) through epigenetic mechanisms determines the development of pathologies with variable severity. For example, our team recently explored the role of microRNAs (miRNAs) targeting and modulating the components of the inflammasome as potential biomarkers in bladder cancer and during therapy. This suggests that the epigenetic control of inflammasome-related genes could represent a potential target for further investigations of molecular mechanisms regulating inflammatory pathways.
Assuntos
Epigênese Genética , Inflamassomos/genética , Animais , Doença/genética , Humanos , Inflamassomos/metabolismo , RNA não Traduzido/genética , RNA não Traduzido/metabolismoRESUMO
Gestational diabetes mellitus (GDM) is a condition of impaired glucose tolerance occurring in 1-14% of all pregnancies. This wide range reflects pathological involvement of single nucleotide polymorphisms (SNPs) and maternal weight as risk factors. This study evaluated the association of genetic component and maternal factors to identify women with higher risk of developing GDM. About 240 pregnant women characterized by negative Oral Glucose Tolerance Test (-OGTT) and 38 with positive OGGT (+OGTT) were enrolled. SNPs for ENPP1, NRF1, VEGFA, CEBPA, and PIK3R1 were analyzed by SNP genotyping. An association study was performed and differences in genotype and allele frequencies between cases and controls were analyzed by χ(2) test. +OGTT was associated to high values of pre-gestational body mass index (BMI) and age. SNP for ENPP1 gene was associated to +OGTT, while genetic variants for other genes did not correlate to GDM. ENPP1 homozygous for A allele and heterozygous showed altered frequencies in +OGTT when compared with -OGTT. Association of both pre-gestational BMI and age with AA homozygous genotype increased significantly the risk to +OGTT. Our results demonstrate that correlation of age and pre-gestational BMI with homozygous for A allele increased significantly the risk of impaired glucose tolerance and GDM.
Assuntos
Índice de Massa Corporal , Diabetes Gestacional/genética , Predisposição Genética para Doença , Diester Fosfórico Hidrolases/genética , Polimorfismo de Nucleotídeo Único , Pirofosfatases/genética , Adolescente , Adulto , Fatores Etários , Alelos , Glicemia/genética , Feminino , Frequência do Gene , Estudos de Associação Genética , Genótipo , Teste de Tolerância a Glucose , Humanos , Pessoa de Meia-Idade , Gravidez , Adulto JovemRESUMO
Alzheimer's disease (AD) is the most frequent cause of dementia worldwide and represents one of the leading factors for severe disability in older persons. Although its etiology is not fully known yet, AD may develop due to multiple factors, including inflammation and oxidative stress, conditions where microRNAs (miRNAs) seem to play a pivotal role as a molecular switch. All these aspects may be modulated by nutritional factors. Among them, vitamin E has been widely studied in AD, given the plausibility of its various biological functions in influencing neurodegeneration. From a cohort of old-aged people, we measured eight vitamin E forms (tocopherols and tocotrienols), thirty cytokines/chemokines, and thirteen exosome-extracted miRNAs in plasma of subjects suffering from subjects affected by AD and age-matched healthy controls (HC). The sample population included 80 subjects (40 AD and 40 HC) with a mean age of 77.6 ± 3.8 years, mostly women (45; 56.2%). Of the vitamin E forms, only α-tocopherol differed between groups, with significantly lower levels in AD. Regarding the examined inflammatory molecules, G-CSF, GM-CSF, INF-α2, IL-3, and IL-8 were significantly higher and IL-17 lower in AD than HC. Among all miRNAs examined, AD showed downregulation of miR-9, miR-21, miR29-b, miR-122, and miR-132 compared to controls. MiR-122 positively and significantly correlated with some inflammatory molecules (GM-CSF, INF-α2, IL-1α, IL-8, and MIP-1ß) as well as with α-tocopherol even after correction for age and gender. A final binary logistic regression analysis showed that α-tocopherol serum levels were associated with a higher AD probability and partially mediated by miR-122. Our results suggest an interplay between α-tocopherol, inflammatory molecules, and microRNAs in AD, where miR-122 may be a good candidate as modulating factor.
Assuntos
Doença de Alzheimer , MicroRNAs , Vitamina E , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , alfa-Tocoferol , Doença de Alzheimer/epidemiologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos , Inflamação , Interleucina-8 , MicroRNAs/genéticaRESUMO
Trace metals can be divided into two subgroups considering their pathophysiological effects: the first consists of microelements essential for life (arsenic, cobalt, chromium, copper, fluorine, iron, iodine, manganese, molybdenum, nickel, selenium, silicon, tin, vanadium and zinc), implicated in important metabolic processes; the second includes toxic microelements, such as cadmium (Cd), mercury (Hg), chromium (Cr), and lead (Pb) for living organisms, even at low concentrations. These metals contribute to serious consequences for human health, including male infertility. Studies performed in several in vitro and in vivo models revealed that environmental exposure to toxic pollutants, as heavy metals, negatively affects human male fertility. Stem cells, due to their ability to self-renew and differentiate in several cell types, have been proposed as a useful tool in assisted reproductive technology, permitting the spermatogenesis recovery in patients with irreversible infertility. Considering the effects of heavy metals on male fertility and, from a demographic point of view, the decreased fertility ratio, further strategies are required to maintain a sustainable turn-over of 2 children for woman. We discuss here the findings on the biological effects of heavy metal pollution in the male fertility and underline the related socioeconomic impact on population demography.
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Mercúrio , Metais Pesados , Oligoelementos , Criança , Feminino , Humanos , Masculino , Cromo , Fertilidade , Metais Pesados/toxicidade , Fatores SocioeconômicosRESUMO
Introduction: Among substances released into the environment by anthropogenic activities, the heavy metal cadmium (Cd) is known to induce severe testicular injury causing male subfertility/infertility. Zinc (Zn) is another heavy metal that, unlike Cd, is physiologically present in the testis, being essential for spermatogenesis. We aimed to examine the possibility that 50 µM ZnCl2 could counteract the toxic effects induced by Cd in an in vitro model of porcine prepubertal Sertoli cells (SCs) exposed to both subtoxic (5 µM) and toxic (10 µM) concentrations of CdCl2 for 48 h. Materials and Methods: Apoptosis, cell cycle, and cell functionality were assessed. The gene expression of Nrf2 and its downstream antioxidant enzymes, ERK1/2, and AKT kinase signaling pathways were evaluated. Materials and Results: We found that Zn, in co-treatment with subtoxic and toxic Cd concentration, increased the number of metabolically active SCs compared to Cd exposure alone but restored SC functionality only in co-treatment with subtoxic Cd concentration with respect to subtoxic Cd alone. Exposure of Cd disrupted cell cycle in SCs, and Zn co-treatment was not able to counteract this effect. Cd alone induced SC death through apoptosis and necrosis in a dose-dependent manner, and co-treatment with Zn increased the pro-apoptotic effect of Cd. Subtoxic and toxic Cd exposures activated the Nrf2 signaling pathway by increasing gene expression of Nrf2 and its downstream genes (SOD, HO-1, and GSHPx). Zn co-treatment with subtoxic Cd attenuated upregulation on the Nrf2 system, while with toxic Cd, the effect was more erratic. Studying ERK1/2 and AKT pathways as a target, we found that the phosphorylation ratio of p-ERK1/2 and p-AKT was upregulated by both subtoxic and toxic Cd exposure alone and in co-treatment with Zn. Discussion: Our results suggest that Zn could counteract Cd effects by increasing the number of metabolically active SCs, fully or partially restoring their functionality by modulating Nrf2, ERK1/2, and AKT pathways. Our SC model could be useful to study the effects of early Cd exposure on immature testis, evaluating the possible protective effects of Zn.
Assuntos
Cádmio , Zinco , Masculino , Animais , Suínos , Cádmio/toxicidade , Zinco/metabolismo , Células de Sertoli/metabolismo , Fator 2 Relacionado a NF-E2/metabolismo , Estresse Oxidativo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de SinaisRESUMO
The environmental light/dark cycle has left its mark on the body's physiological functions to condition not only our inner biology, but also the interaction with external cues. In this scenario, the circadian regulation of the immune response has emerged as a critical factor in defining the host-pathogen interaction and the identification of the underlying circuitry represents a prerequisite for the development of circadian-based therapeutic strategies. The possibility to track down the circadian regulation of the immune response to a metabolic pathway would represent a unique opportunity in this direction. Herein, we show that the metabolism of the essential amino acid tryptophan, involved in the regulation of fundamental processes in mammals, is regulated in a circadian manner in both murine and human cells and in mouse tissues. By resorting to a murine model of pulmonary infection with the opportunistic fungus Aspergillus fumigatus, we showed that the circadian oscillation in the lung of the tryptophan-degrading enzyme indoleamine 2,3-dioxygenase (IDO)1, generating the immunoregulatory kynurenine, resulted in diurnal changes in the immune response and the outcome of fungal infection. In addition, the circadian regulation of IDO1 drives such diurnal changes in a pre-clinical model of cystic fibrosis (CF), an autosomal recessive disease characterized by progressive lung function decline and recurrent infections, thus acquiring considerable clinical relevance. Our results demonstrate that the circadian rhythm at the intersection between metabolism and immune response underlies the diurnal changes in host-fungal interaction, thus paving the way for a circadian-based antimicrobial therapy.
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Introduction: Aims of this study were to evaluate the functional outcomes of a vaginal wall sling technique in patients with stress urinary incontinence at 20 years after surgery and to evaluate the patient's satisfaction after the surgical procedure. Material and Methods: This was a prospective single-center study on patients with stress urinary incontinence who underwent in situ vaginal sling surgery. Presurgery evaluation included history, pelvic examination, and urodynamic test. All patients completed Urogenital Distress Inventory-6 (UDI-6) questionnaire. They underwent checkups at 1, 3, 6, and 12 months postoperatively and then annually. The sling was created by making a rectangle (15-20â×â25 mm) on the anterior vaginal wall and it was reinforced by one roll of Marlex mesh on each side of the sling. The sutures were passed through the vagina at the suprapubic level after suprapubic incision, above the rectus fascia and tied without excessive tension. Results: From May 1996 to May 2002, 40 women underwent vaginal wall sling surgery for stress urinary incontinence. Last visit was performed on 20 women between March 2020 and April 2020. Median follow-up was 251.3 months (20.9 years) (rangeâ=â204.3-285.4 months). The success rate after 5 years of surgical procedure was 80%; over 5 years, the objective cure rate was 45%. Considering only the group of 13 patients with pure stress urinary incontinence, the objective cure rate decreased to 38%, in particular 7 years after surgery. Women who did not resolve their urinary incontinence needed to undergo a new treatment. At over 5 years after surgery, there was an increase in urgency (pâ=â0.001) and voiding symptoms (pâ=â0.008) and urgency urinary incontinence (UUI) (pâ=â0.04). Ninety-five percent were very much worse or much worse according to the Patient Global Impression of Improvement (PGI-I) scale. Conclusion: The in situ vaginal wall sling does not guarantee good long-term functional outcomes in women with stress urinary incontinence.
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Catheter-associated infections in bladder cancer patients, following radical cystectomy or ureterocutaneostomy, are very frequent, and the development of antibiotic resistance poses great challenges for treating biofilm-based infections. Here, we characterized bacterial communities from catheters of patients who had undergone radical cystectomy for muscle-invasive bladder cancer. We evaluated the efficacy of conventional antibiotics, alone or combined with the human ApoB-derived antimicrobial peptide r(P)ApoBLAla, to treat ureteral catheter-colonizing bacterial communities on clinically isolated bacteria. Microbial communities adhering to indwelling catheters were collected during the patients' regular catheter change schedules (28 days) and extracted within 48 h. Living bacteria were characterized using selective media and biochemical assays. Biofilm growth and novel antimicrobial strategies were analyzed using confocal laser scanning microscopy. Statistical analyses confirmed the relevance of the biofilm reduction induced by conventional antibiotics (fosfomycin, ceftriaxone, ciprofloxacin, gentamicin, and tetracycline) and a well-characterized human antimicrobial peptide r(P)ApoBLAla (1:20 ratio, respectively). Catheters showed polymicrobial communities, with Enterobactericiae and Proteus isolates predominating. In all samples, we recorded a meaningful reduction in biofilms, in both biomass and thickness, upon treatment with the antimicrobial peptide r(P)ApoBLAla in combination with low concentrations of conventional antibiotics. The results suggest that combinations of conventional antibiotics and human antimicrobial peptides might synergistically counteract biofilm growth on ureteral catheters, suggesting novel avenues for preventing catheter-associated infections in patients who have undergone radical cystectomy and ureterocutaneostomy.
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Breast cancer (BC) is the most common cancer with the highest frequency of death among women. BC is highly heterogenic at the genetic, biological, and clinical level. Despite the significant improvements in diagnosis and treatments of BC, the high rate of cancer recurrence and resistance to treatment remains a major challenge in clinical practice. This issue is particularly relevant in Triple-Negative Breast Cancer (TNBC) subtype, for which chemotherapy remains the main standard therapeutic approach. Here, we observed that BC cells, belonging to different subtypes, including the TNBC, display an increased expression of Cockayne Syndrome group A (CSA) protein, which is involved in multiple functions such as DNA repair, transcription, mitochondrial homeostasis, and cell division and that recently was found to confer cell robustness when it is up-regulated. We demonstrated that CSA ablation by AntiSense Oligonucleotides (ASOs) drastically impairs tumorigenicity of BC cells by hampering their survival and proliferative capabilities without damaging normal cells. Moreover, suppression of CSA dramatically sensitizes BC cells to platinum and taxane derivatives, which are commonly used for BC first-line therapy, even at very low doses not harmful to normal cells. Finally, CSA ablation restores drug sensitivity in oxaliplatin-resistant cells. Based on these results, we conclude that CSA might be a very attractive target for the development of more effective anticancer therapies.
RESUMO
Fertility preservation for prepubertal male patients undergoing gonadotoxic therapies, potentially depleting spermatogonial cells, is an expanding necessity, yet most of the feasible options are still in the experimental phase. We present our experience and a summary of current and novel possibilities regarding the different strategies to protect or restore fertility in young male patients, before proceeding with chemotherapy or radiotherapy for malignances or other diseases. Adult oncological patients should always be counselled to cryopreserve the semen before starting treatment, however this approach is not suitable for prepubertal boys, who aren't capable to produce sperm yet. Fortunately, since the survival rate of pediatric cancer patients has skyrocketed in the last decade and it's over 84%, safeguarding their future fertility is becoming a major concern for reproductive medicine. Surgical and medical approaches to personalize treatment or protect the gonads could be a valid first step to take. Testicular tissue autologous grafting or xenografting, and spermatogonial stem cells (SSCs) transplantation, are the main experimental options available, but spermatogenesis in vitro is becoming an intriguing alternative. All of these methods feature both strong and weak prospects. There is also relevant controversy regarding the type of testicular material to preserve and the cryopreservation methods. Since transplanted cells are bound to survive based on SSCs number, many ways to enrich their population in cultures have been proposed, as well as different sites of injection inside the testis. Testicular tissue graft has been experimented on mice, rabbits, rhesus macaques and porcine, allowing the birth of live offspring after performing intracytoplasmic sperm injection (ICSI), however it has never been performed on human males yet. In vitro spermatogenesis remains a mirage, although many steps in the right direction have been performed. The manufacturing of 3D scaffolds and artificial spermatogenetic niche, providing support to stem cells in cultures, seems like the best way to further advance in this field.
Assuntos
Preservação da Fertilidade , Neoplasias , Animais , Humanos , Macaca mulatta , Masculino , Camundongos , Neoplasias/terapia , Coelhos , Sêmen , Suínos , TestículoRESUMO
BACKGROUND: The nursing role significantly changed following reforms in the nurse training process. Nowadays, nurses are increasingly trained to promote and improve the quality of clinical practice and to provide support in the assistance of patients and communities. Opportunities and threats are emerging as a consequence of the introduction of new disruptive technologies in public health, which requires the health care staff to develop new digital skills. OBJECTIVE: The aim of this paper is to review and define the role of nurses and the skills they are asked to master in terms of new methodological approaches and digital knowledge in a continuously evolving health care scenario that relies increasingly more on technology and digital solutions. METHODS: This scoping review was conducted using a thematic summary of previous studies. Authors collected publications through a cross-database search (PubMed, Web of Science, Google Scholar) related to new telemedicine approaches impacting the nurses' role, considering the time span of 2011-2021 and therefore including experiences and publications related to the first phase of the COVID-19 pandemic. RESULTS: The assessment was completed between April and July 2021. After a cross-database search, authors reviewed a selection of 60 studies. The results obtained were organized into 5 emerging macro areas: (1) leadership (nurses are expected to show leadership capabilities when introducing new technologies in health care practices, considering their pivotal role in coordinating various professional figures and the patient), (2) soft skills (new communication skills, adaptiveness, and problem solving are needed to adapt the interaction to the level of digital skills and digital knowledge of the patient), (3) training (specific subjects need to be added to nursing training to boost the adoption of new communication and technological skills, enabling health care professionals to largely and effectively use new digital tools), (4) remote management of COVID-19 or chronic patients during the pandemic (a role that has proved to be fundamental is the community and family nurse and health care systems are adopting novel assistance models to support patients at home and to enable decentralization of services from hospitals to the territory), and (5) management of interpersonal relationships with patients through telemedicine (a person-centered approach with an open and sensitive attitude seems to be even more important in the framework of telemedicine where a face-to-face session is not possible and therefore nonverbal indicators are more problematic to be noticed). CONCLUSIONS: Further advancing nurses' readiness in adopting telemedicine requires an integrated approach, including combination of technical knowledge, management abilities, soft skills, and communication skills. This scoping review provides a wide-ranging and general-albeit valuable-starting point to identify these core competences and better understand their implications in terms of present and future health care professionals' roles.