Differential regulation of miR-146a/FAS and miR-21/FASLG axes in autoimmune lymphoproliferative syndrome due to FAS mutation (ALPS-FAS).

Most cases of autoimmune lymphoproliferative syndrome (ALPS) have an inherited genetic defect involving apoptosis-related genes of the FAS pathway. MicroRNAs (miRNAs) are a class of small non-coding regulatory RNAs playing a role in the control of gene expression. This is the first report on miRNAs in ALPS patients. We studied a mother and son carrying the same FAS cell surface death receptor (FAS) mutation, but with only the son manifesting the signs and symptoms of ALPS-FAS. The aim was to analyse, by reverse transcription-quantitative polymerase chain reaction (RT-qPCR), the peripheral blood mononuclear cells (PBMC) relative expression of miR-146a and miR-21, including their passenger strands and respective targets (FAS and FASLG). In comparison with healthy matched control individuals, miR-21-3p was over-expressed significantly (P = 0·0313) in the son, with no significant change in the expression of miR-146a, miR-146a-3p and miR-21. In contrast, the mother had a slight under-expression of the miR-146a pair and miR-21-3p (P = 0·0625). Regarding the miRNA targets, FAS was up-regulated markedly for the mother (P = 0·0078), but down-regulated for the son (P = 0·0625), while FASLG did not have any significant alteration. Taken together, our finding clearly suggests a role of the miR-146a/FAS axis in ALPS-FAS variable expressivity in which FAS haploinsufficiency seems to be compensated only in the mother who had the miR-146a pair down-regulated. As only the son had the major clinical manifestations of ALPS-FAS, miR-21-3p should be investigated as playing a critical role in ALPS physiopathology, including the development of lymphoma.

Apoptosis through Death Receptors in Temporal Lobe Epilepsy-Associated Hippocampal Sclerosis.

Seizure models have demonstrated that neuroinflammation and neurodegeneration are preponderant characteristics of epilepsy. Considering the lack of clinical studies, our aim is to investigate the extrinsic pathway of apoptosis in pharmacoresistant temporal lobe epilepsy (TLE) associated with hippocampal sclerosis (HS) patients, TLE(HS). By a specific death receptor-mediated apoptosis array plate, 31 upregulated targets were revealed in the sclerotic hippocampus from TLE(HS) patients. Amongst them are the encoding genes for ligands (FASLG, TNF, and TNFSF10) and death receptors (FAS, TNFRSF1A, TNFRSF10A, and TNFRSF10B). In addition, we evaluated the hippocampal relative mRNA expression of the two TNF receptors, TNFRSF1A and TNFRSF1B, in patients, being both upregulated (n = 14; P < 0.01 and P < 0.04, resp.) when compared to the post mortem control group (n = 4). Our results have clearly suggested that three different death receptor apoptotic systems may be associated with the maintenance and progression of TLE-associated HS: (1) TNF-TNFRSF1A, (2) FASLG-FAS, and (3) TNFSF10-TNFRSF10A/B. Their effects on epilepsy are still scarcely comprehended. Our study points out to TNF and TNF receptor superfamily pathways as important targets for pharmacological studies regarding the benefits of an anti-inflammatory therapy in these patients.

Hippocampal gene expression dysregulation of Klotho, nuclear factor kappa B and tumor necrosis factor in temporal lobe epilepsy patients.

BACKGROUND: Previous research in animal seizure models indicates that the pleiotropic cytokine TNF is an important effector/mediator of neuroinflammation and cell death. Recently, it has been demonstrated that TNF downregulates Klotho (KL) through the nuclear factor kappa B (NFkB) system in animal models of chronic kidney disease and colitis. KL function in the brain is unclear, although Klotho knockout (Kl-/-) mice exhibit neural degeneration and a reduction of hippocampal synapses. Our aim was to verify if the triad KL-NFKB1-TNF is also dysregulated in temporal lobe epilepsy associated with hippocampal sclerosis (TLE(HS)) patients. FINDINGS: We evaluated TNF, NFKB1 and KL relative mRNA expression levels by reverse transcription quantitative PCR (RT-qPCR) in resected hippocampal tissue samples from 14 TLE(HS) patients and compared them to five post mortem controls. Four reference genes were used: GAPDH, HPRT1, ENO2 and TBP. We found that TNF expression was dramatically upregulated in TLE(HS) patients (P <0.005). NFKB1 expression was also increased (P <0.03) while KL was significantly downregulated (P <0.03) in TLE(HS) patients. Hippocampal KL expression had an inverse correlation with NFKB1 and TNF. CONCLUSIONS: Our data suggest that, similar to other inflammatory diseases, TNF downregulates KL through NFkB in TLE(HS) patients. The remarkable TNF upregulation in patients is a strong indication of hippocampal chronic inflammation. Our finding of hippocampal KL downregulation has wide implications not only for TLE(HS) but also for other neuronal disorders related to neurodegeneration associated with inflammation.

Expressão hipocampal de genes envolvidos em vias de apoptose em pacientes com epilepsia do lobo temporal / Hippocampal expression of genes in apoptotic pathways in temporal lobe epilepsy patients

A epilepsia do lobo temporal associada à esclerose hipocampal [ELT(EH)] é o tipo mais comum de epilepsia focal que causa crises refratárias. A morte neuronal na EH pode ser desencadeada por danos excitotóxicos e citocinas específicas. Pesquisas em modelos experimentais de crises convulsivas ressaltaram a citocina pleiotrópica fator de necrose tumoral (TNF) como um importante efetor/mediador de neuroinflamação e morte celular. Além disso, esses modelos sugeriram que o TNF possa ter uma ação dicotômica por meio de seus dois receptores: ativação da morte celular programada (via TNFRSF1A) ou atuação na sobrevivência celular (via TNFRSF1B), através do fator nuclear kappa B (NFkB). Klotho (KL), originalmente identificada como uma proteína antienvelhecimento, tem se destacado como um importante hormônio regulador de cálcio e fósforo. Sua função cerebral é desconhecida; porém, camundongos knockout para Kl apresentam características que remetem ao envelhecimento humano, com neurodegeneração e redução de sinapses no hipocampo. Em modelos de doença renal crônica e colite, foi comprovado que o TNF inibe KL através do NFkB. Nosso objetivo é identificar alvos críticos na epileptogênese e na fisiopatologia molecular da ELT(EH). Avaliamos a expressão relativa do RNAm de cinco genes-alvo: TNF, TNFRSF1A, TNFRSF1B, NFKB1 e KL. A expressão gênica foi avaliada em amostras de tecido hipocampal de 14 pacientes com ELT(EH) e comparadas com cinco amostras de controles post mortem. Além disso, ambos os receptores do TNF foram analisados nas amostras hipocampais por imuno-histoquímica. Todos os cinco genes avaliados apresentaram expressão significantemente alterada nos pacientes com ELT(EH) (P<0,05). A expressão de ambos os receptores foi constatada nos tecidos dos pacientes. Este é o primeiro estudo a relacionar KL e epilepsia. Nossos dados reforçam o componente inflamatório da EH e sugerem que o TNF possa inibir a expressão de KL no hipocampo dos pacientes.

Temporal lobe epilepsy associated with hippocampal sclerosis [TLE(HS)] is the most common form of focal epilepsy that causes refractory seizures. Neuronal death in HS can be triggered by excitotoxic damage and specific cytokines. Previous research in seizure models indicates that the pleiotropic cytokine tumor necrosis factor (TNF) as an important effector/mediator of neuroinflammation and cell death. Through its two receptors, TNF can play a dichotomous role in animal seizures: programmed cell death activation (via TNFRSF1A) or cell survival actuation (via TNFRSF1B), through the nuclear factor kappa B (NFkB) activation. Klotho (KL), originally identified as an antiaging protein, is emerging as an important calciophosphoregulatory hormone. Its cerebral function is unclear; however, the Kl knockout mouse exhibits a phenotype resembling human aging presenting neural degeneration and a reduction of synapses in the hippocampus. Studies have demonstrated that TNF downregulates KL through NFkB in animal models of chronic kidney disease and colitis. Our aim is to identify critical targets in epileptogenesis to clarify the molecular pathophysiology in TLE(HS). We evaluated the relative mRNA expression of five target genes: TNF, TNFRSF1A, TNFRSF1B, NFKB1 and KL. Gene expression was performed in resected hippocampal tissue samples from 14 TLE(HS) patients and compared to five post mortem controls. Moreover, an immunohistochemistry assay was done to verify the activation of both TNF receptors in patient and control tissues. We found that all target genes were differentially regulated in the TLE(HS) patients (P<0.05). Both TNF receptors were clearly activated in patient's tissues. This is the first study relating KL to epilepsy. Our data corroborates the prominent role of inflammation in HS and suggests that TNF might affect KL expression in hippocampus.