👤 John R Raymond

Mitochondrial positioning supports localized energy and signaling requirements. Miro1 is necessary for attachment of mitochondria to microtubule motor proteins for trafficking. When Miro1 is deleted (Miro1-/-) from mouse embryonic fibroblasts (MEFs), mitochondria become sequestered to the perinuclear space, disrupting subcellular signaling gradients. Here, we show that Miro1-/- MEFs grow slower than Miro1+/+ and Miro1-/- MEFs stably re-expressing a Myc-Miro1 plasmid. Miro1-/- MEFs have a decreased percentage of cells in G1 and increased percentage of cells in S phase. We conducted the first ever RNA sequencing experiment dependent upon Miro1 expression and found differentially expressed genes related to MAPK signaling, cell proliferation and migration. ERK1 and ERK2 (ERK1/2, also known as MAPK3 and MAPK1, respectively) phosphorylation is elevated both spatially and temporally following serum stimulation in Miro1-/- MEFs, whereas the expression levels and oxidation of the dual specificity phosphatases (DUSP1-DUSP6) is unchanged. Finally, we found the oxidation status of ERK1/2 is increased in Miro1-/- MEFs compared to that seen in Miro1+/+ and Myc-Miro1 MEFs. These results highlight transcriptional control based off Miro1 expression and demonstrate the dynamic regulation of ERK1/2 upon deletion of Miro1 which might support the observed cell cycle and proliferation defects. Show less

The early phase of drug development relies on the examination of the efficacy and safety of therapeutic agents in animal models. Due to their close genetic and physiological relation to humans, cynomolgus monkeys ( Show less

Previous reports of distal deletions in chromosome 10q in patients have described distinct facial features combined with other neurodevelopmental abnormalities, including intellectual disability. However, the association of interstitial deletions in chromosome 10q with global developmental delay, musculoskeletal abnormalities, and dysmorphic features has not been previously reported. Genetic testing using whole exome sequencing (WES) was performed on three patients with neurodevelopmental delay, musculoskeletal abnormalities and dysmorphic features. Sequencing reads were aligned to the human genome build GRCh37/UCSC hg19 and analysed for both sequence and copy number variants. WES identified similar interstitial deletions in the 10q21.1q21.3 locus in all three patients. The deleted region includes online Mendelian inheritance in man (OMIM)-annotated genes with clinical significance, such as This is the first report associating interstitial deletions in the 10q21.1q21.3 locus with neurodevelopmental delay, musculoskeletal abnormalities and dysmorphic features. Our findings highlight the clinical significance of this deleted region and suggest possible mechanisms underlying the observed pathological phenotypes. Show less

Mutations in genes traditionally associated with syndromic retinal disease are increasingly found to cause nonsyndromic inherited retinal degenerations. Mutations in CLN3 are classically associated with juvenile neuronal ceroid lipofuscinosis, a rare neurodegenerative disease with early retinal degeneration and progressive neurologic deterioration, but have recently also been identified in patients with nonsyndromic inherited retinal degenerations. To our knowledge, detailed clinical characterization of such cases has yet to be reported. To provide detailed clinical, electrophysiologic, structural, and molecular genetic findings in nonsyndromic inherited retinal degenerations associated with CLN3 mutations. A multi-institutional case series of 10 patients who presented with isolated nonsyndromic retinal disease and mutations in CLN3. Patient ages ranged from 16 to 70 years; duration of follow-up ranged from 3 to 29 years. Longitudinal clinical evaluation, including full ophthalmic examination, multimodal retinal imaging, perimetry, and electrophysiology. Molecular analyses were performed using whole-genome sequencing or whole-exome sequencing. Electron microscopy studies of peripheral lymphocytes and CLN3 transcript analysis with polymerase chain reaction amplification were performed in a subset of patients. There were 7 females and 3 males in this case series, with a mean (range) age at last review of 37.1 (16-70) years. Of the 10 patients, 4 had a progressive late-onset rod-cone dystrophy, with a mean (range) age at onset of 29.7 (20-40) years, and 6 had an earlier onset rod-cone dystrophy, with a mean (range) age at onset of 12.1 (7-17) years. Ophthalmoscopic examination features included macular edema, mild intraretinal pigment migration, and widespread atrophy in advanced disease. Optical coherence tomography imaging demonstrated significant photoreceptor loss except in patients with late-onset disease who had a focal preservation of the ellipsoid zone and outer nuclear layer in the fovea. Electroretinography revealed a rod-cone pattern of dysfunction in 6 patients and were completely undetectable in 2 patients. Six novel CLN3 variants were identified in molecular analyses. This report describes detailed clinical, imaging, and genetic features of CLN3-associated nonsyndromic retinal degeneration. The age at onset and natural progression of retinal disease differs greatly between syndromic and nonsyndromic CLN3 disease, which may be associated with genotypic differences. Show less

In order to identify the potential peripheral signals of appetite and satiety from duodenum, we have performed a transcriptomic study in the mucosa after high-fat (HF) and low-fat (LF) meal ingestion. After fasting, one group of mice was killed and the others were fed ad libitum with HF or LF diet, and killed 30 min, 1 h, and 3 h after the beginning of the meal. The duodenum mucosa was sampled, and the serial analysis of gene expression (SAGE) method was performed. The mRNA regulations were confirmed by real-time PCR. Energy, protein, and fat intakes were higher in the HF than in the LF group. Gene expression profile revealed 118 characterized or partially characterized differentially expressed transcripts. The HF meal delayed the expressions of peptidases compared to the LF groups. Most of mRNAs related to fat absorption, including apolipoprotein A-IV (Apoa4), were decreased in HF1h group, whereas plasma triglyceride (TG) levels were comparable between HF and LF groups. Noteworthy, these downregulations were concomitant to a break in fat intake 1 h after HF meal. At the same time, the HF meal induced transcripts related to cell growth and organization, whereas transcripts involved in cell defense were repressed. Moreover, we have identified fat-responsive transcripts. This study has characterized the molecular responses of duodenum mucosa after HF or LF meal ingestion. Characterization of novel fat-specific candidates whose relations with feeding behavior have never been reported may contribute to the development of new therapeutic targets for appetite and satiety controls. Show less

Continuous modes of renal replacement therapy (CRRT) are increasingly being utilized in the intensive care unit. The removal of cytokines and other inflammatory proteins during ultrafiltration may be responsible for some of the beneficial effects of CRRT. We used proteomic tools to identify proteins found in the ultrafiltrate from a patient with acute renal failure. Identification of these proteins could help elucidate the mechanism(s) of improved outcome with continuous renal replacement therapy. Protein was loaded on a reversed-phase C4 column and eluted with stepwise isocratic flows starting with 0%, 5%, 10%, 25%, and 50% of acetonitrile. Effluent was collected, pooled, desalted, and separated by two-dimensional gel electrophoresis (2DE). Reversed-phase separation improved the resolution and the number of spots seen on the gels. Protein spots were digested with trypsin and spotted onto MALDI plates. Proteins were identified by either peptide mass fingerprinting using a MALDI-TOF mass spectrometer or by peptide sequencing using a MALDI-TOF/TOF tandem mass spectrometer. From 196 spots cut, 47 were identified, representing multiple charge forms of 10 different proteins. Proteins identified were albumin, apolipoprotein A-IV, beta-2-microglobulin, lithostathine, mannose-binding lectin associated serine protease 2 associated protein, plasma retinol-binding protein, transferrin, transthyretin, vitamin D-binding protein and Zn alpha-2 glycoprotein. Continuous renal replacement therapy is frequently used in acutely ill patients with renal failure. Removal of proteins occurs during this process. The physiological significance of this protein removal is unclear. Identification of these proteins will lead to better understanding of the role of protein removal in continuous renal replacement therapy. Show less