Sex-specific Gene Expression in Flupirtine-Treated Cln3Δex7/8 Mouse Brain

Gene expression is a powerful tool to understand structure-function relationships in the nervous system. This study reports global gene expression changes induced by flupirtine in brain of male and female Cln3Δex7/8 mice, exposing potential flupirtine targets at the molecular level. Gene expression analysis of male and female Cln3Δex7/8 mouse brain was determined following oral administration of flupirtine for 14 weeks, using Mouse Genome 430 2.0 array Chips and an Affymetrix platform. Fifty-six genes in males and 79 in females were differentially expressed in flupirtine- versus vehicle-treated Cln3Δex7/8 mouse brain. Flupirtine altered several pathways in Cln3Δex7/8 mouse brain: apoptosis, the complement cascade, NF-kB, and p38α MAPK signaling pathways. Gene-gene network analysis highlighted networks and processes functionally pertinent to flupirtine treatment. These encompassed neurodegeneration, neuro-inflammation, and implicated neurological disorders such as Alzheimer and Parkinson disease. Flupirtine mediates its action in males and females through distinctive actionable targets in the same pathways. This work consolidates the groundwork for considering flupirtine as a treatment option in human CLN3 disease

Exogenous Flupirtine as Potential Treatment for CLN3 Disease

CLN3 disease is a fatal neurodegenerative disorder affecting children. Hallmarks include brain atrophy, accelerated neuronal apoptosis, and ceramide elevation. Treatment regimens are supportive, highlighting the importance of novel, disease-modifying drugs. Flupirtine and its new allyl carbamate derivative (compound 6) confer neuroprotective effects in CLN3-deficient cells. This study lays the groundwork for investigating beneficial effects in Cln3Δex7/8 mice. WT/Cln3Δex7/8 mice received flupirtine/compound 6/vehicle for 14 weeks. Short-term effect of flupirtine or compound 6 was tested using a battery of behavioral testing. For flupirtine, gene expression profiles, astrogliosis, and neuronal cell counts were determined. Flupirtine improved neurobehavioral parameters in open field, pole climbing, and Morris water maze tests in Cln3Δex7/8 mice. Several anti-apoptotic markers and ceramide synthesis/degradation enzymes expression was dysregulated in Cln3Δex7/8 mice. Flupirtine reduced astrogliosis in hippocampus and motor cortex of male and female Cln3Δex7/8 mice. Flupirtine increased neuronal cell counts in male mice. The newly synthesized compound 6 showed promising results in open field and pole climbing. In conclusion, flupirtine improved behavioral, neuropathological and biochemical parameters in Cln3Δex7/8 mice, paving the way for potential therapies for CLN3 disease

Developmental Comparison of Ceramide in Wild-Type and Cln3Δex7/8 Mouse Brains and Sera

CLN3 disease is a neurodevelopmental disease leading to early visual failure, motor decline, and death. CLN3 pathogenesis has been linked to dysregulation of ceramide, a key intracellular messenger impacting various biological functions. Ceramide is upregulated in brains of CLN3 patients and activates apoptosis. Ceramide levels over the lifespan of WT and Cln3Δex7/8 mice were measured using the DGK assay. Ceramide subspecies were determined by LC-MS. Ceramide synthesis enzymes and pre- and post-synaptic mRNA expression was measured in Cln3Δex7/8 and normal mouse brains. Neuronal cell death was established by PARP cleavage and Caspases 3/6/9 and cytochrome C mRNA expression in Cln3Δex7/8 and normal mouse brains. In WT mouse, a ceramide peak was noted at 3 weeks of age. The absence of this peak in Cln3Δex7/8 mice might be related to early disease pathogenesis. Increase of ceramide in Cln3Δex7/8 mouse brain at 24 weeks of age precedes neuronal apoptosis. The correlation between serum and brain ceramide in WT mice, and dysregulation of ceramide in serum and brain of Cln3Δex7/8 mice, and the significant increase in ceramide in Cln3Δex7/8 mouse brains and sera argue for use of easily accessible serum ceramide levels to track response to novel therapies in human CLN3 disease

Wnt/β-Catenin Signaling Pathway Is a Direct Enhancer of Thyroid Transcription Factor-1 in Human Papillary Thyroid Carcinoma Cells

The Wnt/β-catenin signaling pathway is involved in the normal development of thyroid gland, but its disregulation provokes the appearance of several types of cancers, including papillary thyroid carcinomas (PTC) which are the most common thyroid tumours. The follow-up of PTC patients is based on the monitoring of serum thyroglobulin levels which is regulated by the thyroid transcription factor 1 (TTF-1): a tissue-specific transcription factor essential for the differentiation of the thyroid. We investigated whether the Wnt/β-catenin pathway might regulate TTF-1 expression in a human PTC model and examined the molecular mechanisms underlying this regulation. Immunofluorescence analysis, real time RT-PCR and Western blot studies revealed that TTF-1 as well as the major Wnt pathway components are co-expressed in TPC-1 cells and human PTC tumours. Knocking-down the Wnt/β-catenin components by siRNAs inhibited both TTF-1 transcript and protein expression, while mimicking the activation of Wnt signaling by lithium chloride induced TTF-1 gene and protein expression. Functional promoter studies and ChIP analysis showed that the Wnt/β-catenin pathway exerts its effect by means of the binding of β-catenin to TCF/LEF transcription factors on the level of an active TCF/LEF response element at [−798, −792 bp] in TTF-1 promoter. In conclusion, we demonstrated that the Wnt/β-catenin pathway is a direct and forward driver of the TTF-1 expression. The localization of TCF-4 and TTF-1 in the same area of PTC tissues might be of clinical relevance, and justifies further examination of these factors in the papillary thyroid cancers follow-up

Sex differences in gene expression with galactosylceramide treatment in Cln3Δex7/8 mice.

BackgroundCLN3 disease is caused by mutations in the CLN3 gene. The purpose of this study is to discern global expression patterns reflecting therapeutic targets in CLN3 disease.MethodsDifferential gene expression in vehicle-exposed mouse brain was determined after intraperitoneal vehicle/Galactosylceramide (GalCer) injections for 40 weeks with GeneChip Mouse Genome 430 2.0 arrays.ResultsAnalysis identified 66 genes in male and 30 in female brains differentially expressed in GalCer-treated versus vehicle-exposed Cln3Δex7/8 mice. Gene ontology revealed aberrations of biological function including developmental, cellular, and behavioral processes. GalCer treatment altered pathways of long-term potentiation/depression, estrogen signaling, synaptic vesicle cycle, ErbB signaling, and prion diseases in males, but prolactin signaling, selenium compound metabolism and steroid biosynthesis in females. Gene-gene network analysis highlighted networks functionally pertinent to GalCer treatment encompassing motor dysfunction, neurodegeneration, memory disorder, inflammation and astrogliosis in males, and, cataracts, inflammation, astrogliosis, and anxiety in females.ConclusionsThis study sheds light on global expression patterns following GalCer treatment of Cln3Δex7/8 mice. Understanding molecular effects of GalCer on mouse brain gene expression, paves the way for personalized strategies for treating this debilitating disease in humans

Exogenous Flupirtine as Potential Treatment for CLN3 Disease

CLN3 disease is a fatal neurodegenerative disorder affecting children. Hallmarks include brain atrophy, accelerated neuronal apoptosis, and ceramide elevation. Treatment regimens are supportive, highlighting the importance of novel, disease-modifying drugs. Flupirtine and its new allyl carbamate derivative (compound 6) confer neuroprotective effects in CLN3-deficient cells. This study lays the groundwork for investigating beneficial effects in Cln3Δex7/8 mice. WT/Cln3Δex7/8 mice received flupirtine/compound 6/vehicle for 14 weeks. Short-term effect of flupirtine or compound 6 was tested using a battery of behavioral testing. For flupirtine, gene expression profiles, astrogliosis, and neuronal cell counts were determined. Flupirtine improved neurobehavioral parameters in open field, pole climbing, and Morris water maze tests in Cln3Δex7/8 mice. Several anti-apoptotic markers and ceramide synthesis/degradation enzymes expression was dysregulated in Cln3Δex7/8 mice. Flupirtine reduced astrogliosis in hippocampus and motor cortex of male and female Cln3Δex7/8 mice. Flupirtine increased neuronal cell counts in male mice. The newly synthesized compound 6 showed promising results in open field and pole climbing. In conclusion, flupirtine improved behavioral, neuropathological and biochemical parameters in Cln3Δex7/8 mice, paving the way for potential therapies for CLN3 disease

Lithium enhances remyelination of peripheral nerves

Glycogen synthase kinase 3β (GSK3β) inhibitors, especially the mood stabilizer lithium chloride, are also used as neuroprotective or anti-inflammatory agents. We studied the influence of LiCl on the remyelination of peripheral nerves. We showed that the treatment of adult mice with LiCl after facial nerve crush injury stimulated the expression of myelin genes, restored the myelin structure, and accelerated the recovery of whisker movements. LiCl treatment also promoted remyelination of the sciatic nerve after crush. We also demonstrated that peripheral myelin gene MPZ and PMP22 promoter activities, transcripts, and protein levels are stimulated by GSK3β inhibitors (LiCl and SB216763) in Schwann cells as well as in sciatic and facial nerves. LiCl exerts its action in Schwann cells by increasing the amount of β-catenin and provoking its nuclear localization. We showed by ChIP experiments that LiCl treatment drives β-catenin to bind to T-cell factor/lymphoid-enhancer factor response elements identified in myelin genes. Taken together, our findings open perspectives in the treatment of nerve demyelination by administering GSK3β inhibitors such as lithium

A Signature of Four Circulating microRNAs as Potential Biomarkers for Diagnosing Early-Stage Breast Cancer

Breast cancer (BC) is the most predominant type of cancer among women. The aim of this study is to find new biomarkers that can help in early detection of BC, especially for those who are too young to be screened using mammography as per guidelines. Using microRNA microarray, we previously showed dysregulation of 74 microRNAs in tumors from early BC patients as compared with normal adjacent tissues, which we were interested in studying in blood circulation. In this study, we investigated the expression of 12 microRNA (miR-21/miR-155/miR-23a/miR-130a/miR-145/miR-425-5p/miR-139-5p/miR-451/miR-195/miR-125b/miR-100, and miR-182) in the plasma of 41 newly diagnosed Lebanese BC patients with early invasive ductal carcinoma as compared with 32 healthy controls. Total RNA was extracted from plasma, and expression levels of miRNA of interest were measured using RT-qPCR followed by statistical analysis; miR-21, miR-155, miR-23a, miR-130a, miR-145, miR-425-5p, and miR-139-5p were significantly upregulated and miR-451 was significantly downregulated, in the plasma of BC patients as compared with healthy controls. The positively correlated miR-23a, miR-21, and miR-130a had a high diagnostic accuracy (86%). Importantly, the combination of miR-145/miR-425-5p/miR-139-5p/miR-130a scored the highest diagnostic accuracy of 95% with AUC = 0.97 (sensitivity 97% and specificity 91%). MicroRNAs are promising non-invasive diagnostic biomarkers for early-stage BC with the panel of miR-145/miR-425-5p/miR-139-5p/miR-130a having the highest diagnostic accuracy