RESUMO
Diamond Blackfan anemia (DBA) is an inherited bone marrow failure syndrome characterized by congenital anomalies, cancer predisposition and a severe hypo-proliferative anemia. It was the first disease linked to ribosomal dysfunction and >70 % of patients have been identified to have a haploinsufficiency of a ribosomal protein (RP) gene, with RPS19 being the most common mutation. There is significant variability within the disease in terms of phenotype as well as response to therapy suggesting that other genes contribute to the pathophysiology and potential management of this disease. To explore these questions, we performed a genome-wide CRISPR screen in a cellular model of DBA and identified Calbindin 1 (CALB1), a member of the calcium-binding superfamily, as a potential modifier of the disordered erythropoiesis in DBA. We used human derived CD34+ cells cultured in erythroid stimulating media with knockdown of RPS19 as a model for DBA to study the effects of CALB1. We found that knockdown of CALB1 in this DBA model promoted erythroid maturation. We also noted effects of CALB1 knockdown on cell cycle. Taken together, our results reveal CALB1 is a novel regulator of human erythropoiesis and has implications for using CALB1 as a novel therapeutic target in DBA.
Assuntos
Anemia de Diamond-Blackfan , Anemia , Humanos , Anemia de Diamond-Blackfan/genética , Anemia de Diamond-Blackfan/terapia , Eritropoese/genética , Calbindina 1/genética , MutaçãoRESUMO
Rabies virus (RABV) infection leads to a fatal neurological outcome in humans and animals and is associated with major alterations in cellular gene expression. In this study, we describe the effects of RABV infection on the mRNA expression levels of two genes, encoding the Ca2+-binding proteins (Ca-BPs) calbindin D-28K (Calb1) and calretinin (Calb2), in the brains of BALB/c mice. Sixty 4-week-old mice were divided into two test groups and one control group. Mice were inoculated intramuscularly with either a street rabies virus (SRV) strain or a challenge virus standard (CVS-11) strain and sacrificed at 3-day intervals up to day 18 postinfection. A direct fluorescent antibody test (DFAT) was used to verify the presence of RABV antigen in brain tissues, and real-time quantitative PCR (RT-PCR) was used to assess gene expression. Infection with both RABV strains resulted in significant (p < 0.05) increases in Calb1 and Calb2 expression in the test animals when compared with the controls at various time points in the study. Correlation analysis indicated very weak insignificant (p > 0.05) negative and positive relationships, respectively, between Calb1 expression (r = -0.04) and Calb2 expression (r = 0.08) with viral load (CVS-11 strain). Insignificant (p > 0.05) relationships were also observed Calb1 expression (r = -0.28) and Calb2 expression (r = 0.06) and viral load for the SRV strain.The observed alterations in Calb1 and Calb2 expression in this study indicate possible impairments in neuronal Ca2+ buffering and Ca2+ homeostasis as a result of RABV infection and, consequently, possible involvement of calbindin-D28K and calretinin in the neuropathogenesis of rabies.
Assuntos
Encéfalo , Calbindina 1 , Calbindina 2 , Raiva , Animais , Camundongos , Encéfalo/metabolismo , Encéfalo/virologia , Calbindina 2/genética , Raiva/metabolismo , Raiva/patologia , Vírus da Raiva/genética , RNA Mensageiro/genética , Camundongos Endogâmicos BALB C/genética , Calbindina 1/genéticaRESUMO
Generalization is a fundamental cognitive ability of organisms to deal with the uncertainty in real-world situations. Excessive fear generalization and impaired reward generalization are closely related to many psychiatric disorders. However, the neural circuit mechanism for reward generalization and its role in anxiety-like behaviours remain elusive. Here, we found a robust activation of calbindin 1-neurons (Calb 1) in the posterior basolateral amygdala (pBLA), simultaneous with reward generalization to an ambiguous cue after reward conditioning in mice. We identify the infralimbic medial prefrontal cortex (IL) to the pBLACalb1 (Calb 1 neurons in the pBLA) pathway as being involved in reward generalization for the ambiguity. Activating IL-pBLA inputs strengthens reward generalization and reduces chronic unpredictable mild stress-induced anxiety- and depression-like behaviours in a manner dependent on pBLACalb1 neuron activation. These findings suggest that the IL-pBLACalb1 circuit could be a target to promote stress resilience via reward generalization and consequently ameliorate anxiety- and depression-like behaviours.
Assuntos
Ansiedade , Complexo Nuclear Basolateral da Amígdala , Calbindina 1 , Depressão , Neurônios , Córtex Pré-Frontal , Animais , Ansiedade/genética , Ansiedade/metabolismo , Complexo Nuclear Basolateral da Amígdala/metabolismo , Calbindina 1/genética , Calbindina 1/metabolismo , Depressão/genética , Depressão/metabolismo , Camundongos , Neurônios/metabolismo , Neurônios/fisiologia , Córtex Pré-Frontal/metabolismo , Córtex Pré-Frontal/fisiologiaRESUMO
Herein, we aimed to use double-labeling immunofluorescence to describe the expression pattern of Calbindin-D28K (CaBP28K) in the mouse cochlea from late embryonic (E) stages to the adulthood. CaBP28K was expressed in the inner hair cells (IHCs) and the greater epithelial ridge (GER) at E17. In addition, its expression was observed in the interdental cells. On postnatal day 1 (P1), CaBP28K immunoreactivity was observed in the IHCs and outer hair cells (OHCs) and was also specifically expressed in the nucleus and the cytoplasm of spiral ganglion neurons (SGNs). At P8, CaBP28K labeling disappeared from the interdental cells, and the CaBP28K-positive domain within the GER shifted from the entire cytoplasm to only the apical and basal regions. At P14, CaBP28K immunoreactivity was lost from the GER; however, its expression in the IHCs and OHCs, as well as the SGNs, persisted into adulthood. The identification of CaBP28K in the hair cells (HCs) and cuticular plates, as well as SGNs, was confirmed by its colocalization with several markers for Sox2, Myosin VIIa, Phalloidin, and Tuj1. We also detected colocalization with calmodulin in the cytoplasm of both HCs and SGNs. Western blot revealed an increase in CaBP28K postnatal expression in the mouse cochlea.
Assuntos
Calbindina 1/genética , Cóclea/crescimento & desenvolvimento , Neurônios , Gânglio Espiral da Cóclea , Animais , Calbindina 1/análise , Calbindina 1/metabolismo , Imunofluorescência , Imuno-Histoquímica , Camundongos , Neurônios/metabolismoRESUMO
Alzheimer's Disease (AD) is a neurodegenerative disease featured by cognitive impairment. This bioinformatic analysis was used to identify hub genes related to cognitive dysfunction in AD. The gene expression profile GSE48350 in the hippocampus of AD patients aged >70 years was obtained from the Gene Expression Omnibus (GEO) database. A total of 96 differentially expressed genes (DEGs) were identified, and subjected to Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses; a protein-protein interaction (PPI) network was constructed. The DEGs were enriched in synapse-related changes. A protein cluster was teased out of PPI. Furthermore, the cognition ranked the first among all the terms of biological process (BP). Next, 4 of 10 hub genes enriched in cognition were identified. The function of these genes was validated using APP/PS1 mice. Cognitive performance was validated by Morris Water Maze (MWM), and gene expression by RT-qPCR, Cholecystokinin (CCK), Tachykinin precursor 1 (TAC1), Calbindin 1 (CALB1) were downregulated in the hippocampus. These genes can provide new directions in the research of the molecular mechanism of AD.
Assuntos
Doença de Alzheimer , Calbindina 1 , Cognição , Hipocampo/metabolismo , Proteína Serina-Treonina Quinase 2 de Interação com Receptor , Taquicininas , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Animais , Calbindina 1/biossíntese , Calbindina 1/genética , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Transgênicos , Proteína Serina-Treonina Quinase 2 de Interação com Receptor/biossíntese , Proteína Serina-Treonina Quinase 2 de Interação com Receptor/genética , Taquicininas/biossíntese , Taquicininas/genéticaRESUMO
Alzheimer's disease is a progressive, devastating, and irreversible brain disorder that, day by day, destroys memory skills and social behavior. Despite this, the number of known genes suitable for discriminating between AD patients is insufficient. Among the genes potentially involved in the development of AD, there are the chitinase-like proteins (CLPs) CHI3L1, CHI3L2, and CHID1. The genes of the first two have been extensively investigated while, on the contrary, little information is available on CHID1. In this manuscript, we conducted transcriptome meta-analysis on an extensive sample of brains of healthy control subjects (n = 1849) (NDHC) and brains of AD patients (n = 1170) in order to demonstrate CHID1 involvement. Our analysis revealed an inverse correlation between the brain CHID1 expression levels and the age of NDHC subjects. Significant differences were highlighted comparing CHID1 expression of NDHC subjects and AD patients. Exclusive in AD patients, the CHID1 expression levels were correlated positively to calcium-binding adapter molecule 1 (IBA1) levels. Furthermore, both in NDHC and in AD patient's brains, the CHID1 expression levels were directly correlated with calbindin 1 (CALB1) and neurogranin (NRGN). According to brain regions, correlation differences were shown between the expression levels of CHID1 in prefrontal, frontal, occipital, cerebellum, temporal, and limbic system. Sex-related differences were only highlighted in NDHC. CHID1 represents a new chitinase potentially involved in the principal processes underlying Alzheimer's disease.
Assuntos
Doença de Alzheimer/genética , Encéfalo/metabolismo , Calbindina 1/genética , Proteínas de Ligação ao Cálcio/genética , Proteínas de Transporte/genética , Proteínas dos Microfilamentos/genética , Neurogranina/genética , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Encéfalo/patologia , Mapeamento Encefálico , Calbindina 1/metabolismo , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas de Transporte/metabolismo , Estudos de Casos e Controles , Biologia Computacional , Feminino , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Humanos , Masculino , Proteínas dos Microfilamentos/metabolismo , Pessoa de Meia-Idade , Neurogranina/metabolismo , Transdução de Sinais , TranscriptomaRESUMO
Proprioceptive neurons (PNs) are essential for the proper execution of all our movements by providing muscle sensory feedback to the central motor network. Here, using deep single cell RNAseq of adult PNs coupled with virus and genetic tracings, we molecularly identify three main types of PNs (Ia, Ib and II) and find that they segregate into eight distinct subgroups. Our data unveil a highly sophisticated organization of PNs into discrete sensory input channels with distinct spatial distribution, innervation patterns and molecular profiles. Altogether, these features contribute to finely regulate proprioception during complex motor behavior. Moreover, while Ib- and II-PN subtypes are specified around birth, Ia-PN subtypes diversify later in life along with increased motor activity. We also show Ia-PNs plasticity following exercise training, suggesting Ia-PNs are important players in adaptive proprioceptive function in adult mice.
Assuntos
Retroalimentação Sensorial/fisiologia , Gânglios Espinais/metabolismo , Neurônios Motores/metabolismo , Propriocepção/fisiologia , Células Receptoras Sensoriais/metabolismo , Animais , Calbindina 1/genética , Calbindina 1/metabolismo , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteínas Correpressoras/genética , Proteínas Correpressoras/metabolismo , Subunidade alfa 2 de Fator de Ligação ao Core/genética , Subunidade alfa 2 de Fator de Ligação ao Core/metabolismo , Subunidade alfa 3 de Fator de Ligação ao Core/genética , Subunidade alfa 3 de Fator de Ligação ao Core/metabolismo , Gânglios Espinais/citologia , Expressão Gênica , Proteínas com Domínio LIM/genética , Proteínas com Domínio LIM/metabolismo , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios Motores/classificação , Neurônios Motores/citologia , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Condicionamento Físico Animal , Células Receptoras Sensoriais/classificação , Células Receptoras Sensoriais/citologia , Análise de Célula Única , Medula Espinal/citologia , Medula Espinal/metabolismoRESUMO
Calbindin-D28k (CB), a calcium-binding protein, mediates diverse neuronal functions. In this study, adult gerbils were fed a normal diet (ND) or exposed to intermittent fasting (IF) for three months, and were randomly assigned to sham or ischemia operated groups. Ischemic injury was induced by transient forebrain ischemia for 5 min. Short-term memory was examined via passive avoidance test. CB expression was investigated in the Cornu Ammonis 1 (CA1) region of the hippocampus via western blot analysis and immunohistochemistry. Finally, histological analysis was used to assess neuroprotection and gliosis (microgliosis and astrogliosis) in the CA1 region. Short-term memory did not vary significantly between ischemic gerbils with IF and those exposed to ND. CB expression was increased significantly in the CA1 pyramidal neurons of ischemic gerbils with IF compared with that of gerbils fed ND. However, the CB expression was significantly decreased in ischemic gerbils with IF, similarly to that of ischemic gerbils exposed to ND. The CA1 pyramidal neurons were not protected from ischemic injury in both groups, and gliosis (astrogliosis and microgliosis) was gradually increased with time after ischemia. In addition, immunoglobulin G was leaked into the CA1 parenchyma from blood vessels and gradually increased with time after ischemic insult in both groups. Taken together, our study suggests that IF for three months increases CB expression in hippocampal CA1 pyramidal neurons; however, the CA1 pyramidal neurons are not protected from transient forebrain ischemia. This failure in neuroprotection may be attributed to disruption of the blood-brain barrier, which triggers gliosis after ischemic insults.
Assuntos
Calbindina 1/genética , Jejum , Expressão Gênica , Traumatismo por Reperfusão/etiologia , Traumatismo por Reperfusão/metabolismo , Animais , Calbindina 1/imunologia , Morte Celular/genética , Morte Celular/imunologia , Gerbillinae , Gliose/etiologia , Imunoglobulina G/imunologia , Masculino , Neurônios/metabolismo , Neurônios/patologia , Traumatismo por Reperfusão/tratamento farmacológico , Traumatismo por Reperfusão/patologiaRESUMO
Cochlear development is a complex process with precise spatiotemporal patterns. A detailed understanding of this process is important for studies of congenital hearing loss and regenerative medicine. However, much of our understanding of cochlear development is based on rodent models. Animal models that bridge the gap between humans and rodents are needed. In this study, we investigated the development of hearing organs in a small New World monkey species, the common marmoset (Callithrix jacchus). We describe the general stages of cochlear development in comparison with those of humans and mice. Moreover, we examined more than 25 proteins involved in cochlear development and found that expression patterns were generally conserved between rodents and primates. However, several proteins involved in supporting cell processes and neuronal development exhibited interspecific expression differences. Human fetal samples for studies of primate-specific cochlear development are extremely rare, especially for late developmental stages. Our results support the use of the common marmoset as an effective alternative for analyses of primate cochlear development.
Assuntos
Callithrix/genética , Cóclea/metabolismo , Regulação da Expressão Gênica no Desenvolvimento , Modelos Animais , Organogênese/genética , Animais , Aquaporina 4/genética , Aquaporina 4/metabolismo , Calbindina 1/genética , Calbindina 1/metabolismo , Callithrix/embriologia , Callithrix/crescimento & desenvolvimento , Callithrix/metabolismo , Cóclea/anatomia & histologia , Cóclea/citologia , Cóclea/crescimento & desenvolvimento , Sequência Conservada , Inibidor de Quinase Dependente de Ciclina p27/genética , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Embrião de Mamíferos , Fator de Transcrição GATA3/genética , Fator de Transcrição GATA3/metabolismo , Humanos , Proteínas com Homeodomínio LIM/genética , Proteínas com Homeodomínio LIM/metabolismo , Camundongos , Miosina VIIa/genética , Miosina VIIa/metabolismo , Parvalbuminas/genética , Parvalbuminas/metabolismo , Periferinas/genética , Periferinas/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Especificidade da Espécie , Transportadores de Sulfato/genética , Transportadores de Sulfato/metabolismo , Fator de Transcrição Brn-3C/genética , Fator de Transcrição Brn-3C/metabolismo , Tubulina (Proteína)/genética , Tubulina (Proteína)/metabolismoRESUMO
Alzheimer's disease (AD) represents one of the main forms of dementia that afflicts our society. The expression of several genes has been associated with disease development. Despite this, the number of genes known to be capable of discriminating between AD patients according to sex remains deficient. In our study, we performed a transcriptomes meta-analysis on a large court of brains of healthy control subjects (n = 2139) (NDHC) and brains of AD patients (n = 1170). Our aim was to verify the brain expression levels of CHI3L2 and its correlation with genes associated with microglia-mediated neuroinflammation (IBA1), alteration of the blood-brain barrier (PECAM1), and neuronal damage (CALB1). We showed that the CHI3L2, IBA1, PECAM1, and CALB1 expression levels were modulated in the brains of patients with AD compared to NDHC subjects. Furthermore, both in NDHC and in AD patient's brains, the CHI3L2 expression levels were directly correlated with IBA1 and PECAM1 and inversely with CALB1. Additionally, the expression levels of CHI3L2, PECAM1, and CALB1 but not of IBA1 were sex-depended. By stratifying the samples according to age and sex, correlation differences emerged between the expression levels of CHI3L2, IBA1, PECAM1, and CALB1 and the age of NDHC subjects and AD patients. CHI3L2 represents a promising gene potentially involved in the key processes underlying Alzheimer's disease. Its expression in the brains of sex-conditioned AD patients opens up new possible sex therapeutic strategies aimed at controlling imbalance in disease progression.
Assuntos
Doença de Alzheimer/metabolismo , Transcriptoma , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Encéfalo/metabolismo , Calbindina 1/genética , Calbindina 1/metabolismo , Proteínas de Ligação ao Cálcio/genética , Proteínas de Ligação ao Cálcio/metabolismo , Quitinases/genética , Quitinases/metabolismo , Bases de Dados Genéticas , Feminino , Humanos , Masculino , Proteínas dos Microfilamentos/genética , Proteínas dos Microfilamentos/metabolismo , Molécula-1 de Adesão Celular Endotelial a Plaquetas/genética , Molécula-1 de Adesão Celular Endotelial a Plaquetas/metabolismoRESUMO
The basolateral amygdala (BLA) and ventral hippocampal CA1 (vCA1) are cellularly and functionally diverse along their anterior-posterior and superficial-deep axes. Here, we find that anterior BLA (aBLA) and posterior BLA (pBLA) innervate deep-layer calbindin1-negative (Calb1-) and superficial-layer calbindin1-positive neurons (Calb1+) in vCA1, respectively. Photostimulation of pBLA-vCA1 inputs has an anxiolytic effect in mice, promoting approach behaviours during conflict exploratory tasks. By contrast, stimulating aBLA-vCA1 inputs induces anxiety-like behaviour resulting in fewer approaches. During conflict stages of the elevated plus maze task vCA1Calb1+ neurons are preferentially activated at the open-to-closed arm transition, and photostimulation of vCA1Calb1+ neurons at decision-making zones promotes approach with fewer retreats. In the APP/PS1 mouse model of Alzheimer's disease, which shows anxiety-like behaviour, photostimulating the pBLA-vCA1Calb1+ circuit ameliorates the anxiety in a Calb1-dependent manner. These findings suggest the pBLA-vCA1Calb1+ circuit from heterogeneous BLA-vCA1 connections drives approach behaviour to reduce anxiety-like behaviour.
Assuntos
Ansiolíticos/farmacologia , Complexo Nuclear Basolateral da Amígdala/metabolismo , Região CA1 Hipocampal/metabolismo , Calbindina 1/metabolismo , Comportamento de Escolha/fisiologia , Doença de Alzheimer/metabolismo , Animais , Ansiedade , Comportamento Animal , Calbindina 1/genética , Tomada de Decisões , Masculino , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neurônios/fisiologia , ProteômicaRESUMO
Calcineurin inhibitors (CNIs) are immunosuppressive drugs used to prevent graft rejection after organ transplant. Common side effects include renal magnesium wasting and hypomagnesemia, which may contribute to new-onset diabetes mellitus, and hypercalciuria, which may contribute to post-transplant osteoporosis. Previous work suggested that CNIs reduce the abundance of key divalent cation transport proteins, expressed along the distal convoluted tubule, causing renal magnesium and calcium wasting. It has not been clear, however, whether these effects are specific for the distal convoluted tubule, and whether these represent off-target toxic drug effects, or result from inhibition of calcineurin. The CNI tacrolimus can inhibit calcineurin only when it binds with the immunophilin, FKBP12; we previously generated mice in which FKBP12 could be deleted along the nephron, to test whether calcineurin inhibition is involved, these mice are normal at baseline. Here, we confirmed that tacrolimus-treated control mice developed hypomagnesemia and urinary calcium wasting, with decreased protein and mRNA abundance of key magnesium and calcium transport proteins (NCX-1 and Calbindin-D28k ). However, qPCR also showed decreased mRNA expression of NCX-1 and Calbindin-D28k , and TRPM6. In contrast, KS-FKBP12-/- mice treated with tacrolimus were completely protected from these effects. These results indicate that tacrolimus affects calcium and magnesium transport along the distal convoluted tubule and strongly suggests that inhibition of the phosphatase, calcineurin, is directly involved.
Assuntos
Inibidores de Calcineurina/farmacologia , Cálcio/metabolismo , Hipercalciúria/induzido quimicamente , Túbulos Renais Distais/efeitos dos fármacos , Magnésio/metabolismo , Proteína 1A de Ligação a Tacrolimo/genética , Tacrolimo/farmacologia , Desequilíbrio Hidroeletrolítico/induzido quimicamente , Animais , Calbindina 1/efeitos dos fármacos , Calbindina 1/genética , Calbindina 1/metabolismo , Inibidores de Calcineurina/efeitos adversos , Cálcio/urina , Expressão Gênica , Hipercalciúria/metabolismo , Hipercalciúria/urina , Túbulos Renais Distais/metabolismo , Magnésio/urina , Camundongos , Camundongos Knockout , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Trocador de Sódio e Cálcio/efeitos dos fármacos , Trocador de Sódio e Cálcio/genética , Trocador de Sódio e Cálcio/metabolismo , Canais de Cátion TRPM/efeitos dos fármacos , Canais de Cátion TRPM/genética , Canais de Cátion TRPM/metabolismo , Tacrolimo/efeitos adversos , Proteína 1A de Ligação a Tacrolimo/metabolismo , Desequilíbrio Hidroeletrolítico/metabolismo , Desequilíbrio Hidroeletrolítico/urinaRESUMO
Diabetes-associated advanced glycation end-products (AGEs) can increase extracellular matrix (ECM) expression and induce renal fibrosis. Calbindin-D28k, which plays a role in calcium reabsorption in renal distal convoluted tubules, is increased in a diabetic kidney. The role of calbindin-D28k in diabetic nephropathy still remains unclear. Here, calbindin-D28k protein expression was unexpectedly induced in the renal tubules of db/db diabetic mice. AGEs induced the calbindin-D28k expression in human renal proximal tubule cells (HK2), but not in mesangial cells. AGEs induced the expression of fibrotic molecules, ECM proteins, epithelial-mesenchymal transition (EMT) markers, and endoplasmic reticulum (ER) stress-related molecules in HK2 cells, which could be inhibited by a receptor for AGE (RAGE) neutralizing antibody. Calbindin-D28k knockdown by siRNA transfection reduced the cell viability and obviously enhanced the protein expressions of fibrotic factors, EMT markers, and ER stress-related molecules in AGEs-treated HK2 cells. Chemical chaperone 4-Phenylbutyric acid counteracted the AGEs-induced ER stress and ECM and EMT markers expressions. Calbindin-D28k siRNA in vivo delivery could enhance renal fibrosis in db/db diabetic mice. These findings suggest that inducible calbindin-D28k protects against AGEs/RAGE axis-induced ER stress-activated ECM induction and cell injury in renal proximal tubule cells.
Assuntos
Calbindina 1/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Nefropatias Diabéticas/patologia , Produtos Finais de Glicação Avançada/metabolismo , Túbulos Renais Distais/patologia , Animais , Calbindina 1/genética , Diabetes Mellitus Tipo 2/complicações , Nefropatias Diabéticas/etiologia , Nefropatias Diabéticas/metabolismo , Modelos Animais de Doenças , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/patologia , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Fibrose , Técnicas de Silenciamento de Genes , Humanos , Túbulos Renais Distais/efeitos dos fármacos , Masculino , Células Mesangiais/metabolismo , Camundongos , Fenilbutiratos/farmacologia , RNA Interferente Pequeno/metabolismo , Receptor para Produtos Finais de Glicação Avançada/antagonistas & inibidores , Receptor para Produtos Finais de Glicação Avançada/metabolismoRESUMO
Numerous studies have demonstrated the association between senescence and cancer. However, the molecular mechanism regulating senescence in ovarian cancer remains unknown. In the present study, the protein expression level of calbindin 1 (CALB1) in ovarian cancer was examined using western blot and immunohistochemistry. The function of CALB1 in ovarian cancer cells was examined using MTT assay, anchorageindependent growth assay and senescence assay. The molecular mechanisms underlying CALB1 function were investigated using immunoprecipitation and pulldown assays. In the present study, the expression of CALB1 was found to be increased in ovarian cancer. Overexpression of CALB1 promoted the proliferation and colony formation of ovarian cancer cells and inhibited senescence by modulating the expression levels of p21 and p27. Knockdown of CALB1 inhibited the proliferation and colony formation of ovarian cancer cells. Mechanistically, coimmunoprecipitation assays revealed that CALB1 interacts with MDM2 protooncogene (MDM2) and promoted the interaction between p53 and MDM2. Collectively, the present study suggested that CALB1 may act as an oncogene in ovarian cancer by inhibiting the p53 pathway.
Assuntos
Calbindina 1/metabolismo , Senescência Celular , Neoplasias Ovarianas/patologia , Proteínas Proto-Oncogênicas c-mdm2/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Calbindina 1/antagonistas & inibidores , Calbindina 1/genética , Linhagem Celular Tumoral , Proliferação de Células , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Inibidor de Quinase Dependente de Ciclina p27/metabolismo , Feminino , Humanos , Neoplasias Ovarianas/metabolismo , Plasmídeos/genética , Plasmídeos/metabolismo , Ligação Proteica , Interferência de RNA , RNA Interferente Pequeno/metabolismoRESUMO
The subfornical organ (SFO) is forebrain sensory circumventricular organ, characterized by lack of a blood-brain barrier. Neurons of the SFO can detect circulating molecules such as peptide hormones and communicate this information to regulatory centers behind the blood-brain barrier, thus playing a critical role in homeostatic processes including regulation of energy balance, hydromineral balance and cardiovascular control. The SFO contains two subregions defined by neuronal expression of molecular markers: the dorsolateral peripheral or shell SFO (sSFO) neurons express calretinin, and the ventromedial core (cSFO) neurons express calbindin D28K. Neurons from these two subregions project to different locations to subserve different roles in homeostatic regulation. It is unknown whether neurons from these two subregions exhibit unique or identifiable electrophysiological properties. This study used a gold nanoparticle-conjugated RNA fluorescent probe on dissociated SFO neuron cultures and patch clamp electrophysiology to characterize the intrinsic electrophysiological properties of cSFO and sSFO neurons. Our studies revealed that neurons originating from the core region exhibited significantly more action potential bursting, while neurons from non-core regions exhibited more tonic firing neurons, albeit at a higher overall frequency. The difference in activity is correlated with a more depolarized resting membrane potential and a higher density of voltage gated Na+ currents.
Assuntos
Calbindina 1/biossíntese , Fenômenos Eletrofisiológicos/fisiologia , Neurônios/fisiologia , Órgão Subfornical/fisiologia , Animais , Calbindina 1/genética , Células Cultivadas , Expressão Gênica , Masculino , Ratos , Ratos Sprague-DawleyRESUMO
Synthetic glucocorticoids (sGC) are administered to women at risk for pre-term delivery to reduce respiratory distress syndrome in the newborn. The prefrontal cortex (PFC) is important in regulating stress responses and related behaviours and expresses high levels of glucocorticoid receptors (GR). Further, antenatal exposure to sGC results in a hyperactive phenotype in first generation (F1) juvenile male and female offspring, as well as F2 and F3 juvenile females from the paternal lineage. We hypothesized that multiple courses of antenatal sGC modify gene expression in the PFC, that these effects are sex-specific and maintained across multiple generations, and that the gene sets affected relate to modified locomotor activity. We performed RNA sequencing on PFC of F1 juvenile males and females, as well as F2 and F3 juvenile females from the paternal lineage and used regression modelling to relate gene expression and behavior. Antenatal sGC resulted in sex-specific and generation-specific changes in gene expression. Further, the expression of 4 genes (C9orf116, Calb1, Glra3, and Gpr52) explained 20-29% of the observed variability in locomotor activity. Antenatal exposure to sGC profoundly influences the developing PFC; effects are evident across multiple generations and may drive altered behavioural phenotypes.
Assuntos
Glucocorticoides/administração & dosagem , Locomoção/efeitos dos fármacos , Córtex Pré-Frontal/efeitos dos fármacos , Nascimento Prematuro/tratamento farmacológico , Síndrome do Desconforto Respiratório do Recém-Nascido/tratamento farmacológico , Adulto , Calbindina 1/genética , Feminino , Regulação da Expressão Gênica no Desenvolvimento/efeitos dos fármacos , Glucocorticoides/efeitos adversos , Glucocorticoides/síntese química , Humanos , Recém-Nascido , Locomoção/genética , Masculino , Córtex Pré-Frontal/metabolismo , Nascimento Prematuro/genética , Nascimento Prematuro/patologia , Nascimento Prematuro/prevenção & controle , Receptores Acoplados a Proteínas G/genética , Receptores de Glicina/genética , Síndrome do Desconforto Respiratório do Recém-Nascido/genética , Síndrome do Desconforto Respiratório do Recém-Nascido/patologia , Síndrome do Desconforto Respiratório do Recém-Nascido/prevenção & controle , Caracteres Sexuais , Estresse Fisiológico/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacosRESUMO
Delayed hippocampal injury and memory impairments follow neonatal hypoxia-ischemia (HI) despite the use of therapeutic hypothermia (TH). Death of hippocampal pyramidal cells occurs acutely after HI, but characterization of delayed cell death and injury of interneurons (INs) is unknown. We hypothesize that injury of INs after HI is: (i) asynchronous to that of pyramidal cells, (ii) independent of injury severity, and (iii) unresponsive to TH. HI was induced in C57BL6 mice at p10 with unilateral right carotid ligation and 45 min of hypoxia (FiO2 = 0.08). Mice were randomized to normothermia (36 °C, NT) or TH (31 °C) for 4 hr after HI and anesthesia-exposed shams were use as controls. Brains were studied at 24 hr (p11) or 8 days (p18) after HI. Vglut1, GAD65/67, PSD95, parvalbumin (PV) and calbindin-1 (Calb1) were measured. Cell death was assessed using cresyl violet staining and TUNEL assay. Hippocampal atrophy and astroglyosis at p18 were used to assess injury severity and to correlate with number of PV + INs. VGlut1 level decreased by 30% at 24 hr after HI, while GAD65/67 level decreased by â¼50% in forebrain 8 days after HI, a decrease localized in CA1 and CA3. PSD95 levels decreased in forebrain by 65% at 24 hr after HI and remained low 8 days after HI. PV + INs increased in numbers (per mm2 ) and branching between p11 and p18 in sham mice but not in NT and TH mice, resulting in 21-52% fewer PV + INs in injured mice at p18. Calb1 protein and mRNA were also reduced in HI injured mice at p18. At p18, somatodendritic attrition of INs was evident in all injured mice without evidence of cell death. Neither hippocampal atrophy nor astroglyosis correlated with the number of PV + INs at p18. Thus, HI exposure has long lasting effects in the hippocampus impairing the development of the GABAergic system with only partial protection by TH independent of the degree of hippocampal injury. © 2018 Wiley Periodicals, Inc.
Assuntos
Hipocampo/patologia , Hipotermia Induzida/métodos , Hipóxia-Isquemia Encefálica/terapia , Interneurônios/patologia , Animais , Animais Recém-Nascidos , Calbindina 1/genética , Calbindina 1/metabolismo , Modelos Animais de Doenças , Proteína 4 Homóloga a Disks-Large/metabolismo , Lateralidade Funcional , Expressão Gênica/fisiologia , Proteína Glial Fibrilar Ácida/metabolismo , Hipocampo/metabolismo , Hipóxia-Isquemia Encefálica/patologia , Camundongos , Camundongos Endogâmicos C57BL , Parvalbuminas/metabolismo , Tubulina (Proteína)/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Proteínas Vesiculares de Transporte de Glutamato/metabolismoRESUMO
Calbindin (CALB) is well established as immunohistochemical marker for intrinsic primary afferent neurons in the guinea pig gut. Its expression by numerous human enteric neurons has been demonstrated but little is known about particular types of neurons immunoreactive for CALB. Here we investigated small and large intestinal wholemount sets of 26 tumor patients in order to evaluate (1) the proportion of CALB⺠neurons in the total neuron population, (2) the colocalization of CALB with calretinin (CALR), somatostatin (SOM) and vasoactive intestinal peptide (VIP) and (3) the morphology of CALB+ neurons. CALB+ neurons represented a minority of myenteric neurons (small intestine: 31%; large intestine: 25%) and the majority of submucosal neurons (between 72 and 95%). In the submucosa, most CALB⺠neurons co-stained for CALR and VIP (between 69 and 80%) or for SOM (between 20 and 3%). In the myenteric plexus, 85% of CALB+ neurons did not co-stain with the other markers investigated. An unequivocal correlation between CALB reactivity and neuronal morphology was found for myenteric type III neurons in the small intestine: uniaxonal neurons with long, slender and branched dendrites were generally positive for CALB. Since also other neurons displayed occasional CALB reactivity, this protein is not suited as an exclusive marker for type III neurons.
Assuntos
Calbindina 1/metabolismo , Plexo Mientérico/citologia , Neurônios/metabolismo , Plexo Submucoso/citologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Calbindina 1/genética , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Plexo Mientérico/metabolismo , Neurônios/classificação , Somatostatina/genética , Somatostatina/metabolismo , Plexo Submucoso/metabolismo , Peptídeo Intestinal Vasoativo/genética , Peptídeo Intestinal Vasoativo/metabolismoRESUMO
Calcium binding protein calbindin-D28K (CaBP28K) mediates the relationship between vitamin D and calcium, but its mechanism remains unclear during bone formation. The present study reports that maternal CaBP28K levels were positively correlated with paired umbilical cord CaBP28K levels. In addition, CaBP28K levels were positively correlated with the body length, and head and chest circumferences of neonates, but negatively correlated with maternal 25(OH)D3 levels. CaBP28K was also downregulated in MC3T3-E1 osteoblasts when treated with 1,25(OH)2D or VDR overexpression, but was upregulated in the femur of 1α(OH)ase(-/-) mice. Furthermore, it was found CaBP28K may influence cell differentiation and matrix formation through the regulation of DMP1 and the interaction with MMP13 in osteoblasts. This suggests that CaBP28K could be a candidate for the negative role of 1,25(OH)2D/VDR in regulating bone mass.
Assuntos
Calbindina 1/metabolismo , Calcitriol/metabolismo , Proteínas da Matriz Extracelular/metabolismo , Metaloproteinase 13 da Matriz/metabolismo , Osteogênese/fisiologia , Receptores de Calcitriol/metabolismo , Adolescente , Adulto , Animais , Calbindina 1/genética , Linhagem Celular , Proteínas da Matriz Extracelular/genética , Feminino , Humanos , Metaloproteinase 13 da Matriz/genética , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Osteogênese/genética , Adulto JovemRESUMO
Heat stress is a problem in laying hens as it decreases egg quality by decreasing eggshell mineralization. Heat stress alters gene expression, hence our aim was to investigate effects of heat stress on gene expression of ion transport elements involving in uterine mineralization (TRPV6, CALB1, ITPR3, SCNN1G, SLC4A4, KCNJ15, SLC4A9, and CLCN2) by real time quantitative PCR. Forty 23-week-old White Leghorn laying hens were housed in two rooms. The control group (n = 20) was maintained at 21-23 °C, and the heat stress group (n = 20) was exposed to 36-38 °C for 8 weeks. All parameters of egg quality including egg weight, surface area, volume, and eggshell weight, thickness, ash weight, and calcium content were decreased in the heat stress group compared to the control group (by 26.9%, 32.7%, 44.1%, 38.4%, 31.7%, 39.4%, and 11.1%, respectively). Total plasma calcium was decreased by 13.4%. Levels of ITPR3, SLC4A4, and SLC4A9 transcripts in the uterine lining were decreased in the heat stress group compared to the control group (by 61.4%, 66.1%, and 66.1%, respectively). CALB1 transcript level was increased (by 34.2 fold) in the heat stress group of hens compared to controls. TRPV6, SCNN1G, KCNJ15, and CLCN2 transcript levels did not significantly differ between control and heat stress groups of laying hens. It is concluded that the down-expression of ITPR3, SLC4A4, and SLC4A9 genes may impair transportation of Cl-, HCO3-, and Na+ in eggshell mineralization during heat stress. Increased CALB1 gene expression may increase resistance of uterine cells to detrimental effects of heat stress.