Upregulation of Ras/Raf/ERK1/2 signaling and ERK5 in the brain of autistic subjects

Autism is a neurodevelopmental disorder characterized by impairments in social interaction, verbal communication and repetitive behaviors. A number of studies have shown that the Ras/Raf/ERK1/2 (extracellular signal-regulated kinase) signaling pathway plays important roles in the genesis of neural progenitors, learning and memory. Ras/Raf/ERK1/2 and ERK5 have also been shown to have death-promoting apoptotic roles in neural cells. Recent studies have shown a possible association between neural cell death and autism. In addition, two recent studies reported that a deletion of a locus on chromosome 16, which included the mitogen-activated protein kinase 3 (MAPK3) gene that encodes ERK1, is associated with autism. Most recently, our laboratory detected that Ras/Raf/ERK1/2 signaling activities were significantly enhanced in the brain of BTBR mice that model autism, as they exhibit many autism-like behaviors. We thus hypothesized that Ras/Raf/ERK1/2 signaling and ERK5 could be abnormally regulated in the brain of autistic subjects. In this study, we show that the expression of Ras protein was significantly elevated in the frontal cortex of autistic subjects. C-Raf phosphorylation was increased in the frontal cortex, while both C-Raf and A-Raf activities were enhanced in the cerebellum of autistic subjects. We also detected that both the protein expression and activities of ERK1/2 were significantly upregulated in the frontal cortex of autistic subjects, but not in the cerebellum. Furthermore, we showed that ERK5 protein expression is upregulated in both frontal cortex and cerebellum of autistic subjects. These results suggest that the upregulation of Ras/Raf/ERK1/2 signaling and ERK5 activities mainly found in the frontal cortex of autistic subjects may be critically involved in the pathogenesis of autism.     

Loss and re-adaptation of lumbar intervertebral disc water signal intensity after prolonged bedrest

The adaptation and re-adaptation process of the intervertebral disc (IVD) to prolonged bedrest is important for understanding IVD physiology and IVD herniations in astronauts. Little information is available on changes in IVD composition. In this study, 24 male subjects underwent 60-day bedrest and In/Out Phase magnetic resonance imaging sequences were performed to evaluate IVD shape and water signal intensity. Scanning was performed before bedrest (baseline), twice during bedrest, and three, six and twenty-four months after bedrest. Area, signal intensity, average height, and anteroposterior diameter of the lumbar L3/4 and L4/5 IVDs were measured. At the end of bedrest, disc height and area were significantly increased with no change in water signal intensity. After bedrest, we observed reduced IVD signal intensity three months (p=0.004 versus baseline), six months (p=0.003 versus baseline), but not twenty-four months (p=0.25 versus baseline) post-bedrest. At these same time points post-bedrest, IVD height and area remained increased. The reduced lumbar IVD water signal intensity in the first months after bedrest implies a reduction of glycosaminoglycans and/or free water in the IVD. Subsequently, at two years after bedrest, IVD hydration status returned towards pre-bedrest levels, suggesting a gradual, but slow, re-adaptation process of the IVD after prolonged bedrest.     

Association between Rare Variants in AP4E1, a Component of Intracellular Trafficking,and Persistent Stuttering

Stuttering is a common, highly heritable neurodevelopmental disorder characterized by deficits in the volitional control of speech. Whole-exome sequencing identified two heterozygous AP4E1 coding variants, c.1549G>A (p.Val517Ile) and c.2401G>A (p.Glu801Lys), that co-segregate with persistent developmental stuttering in a large Cameroonian family, and we observed the same two variants in unrelated Cameroonians with persistent stuttering. We found 23 other rare variants, including predicted loss-of-function variants, in AP4E1 in unrelated stuttering individuals in Cameroon, Pakistan, and North America. The rate of rare variants in AP4E1 was significantly higher in unrelated Pakistani and Cameroonian stuttering individuals than in population-matched control individuals, and coding variants in this gene are exceptionally rare in the general sub-Saharan West African, South Asian, and North American populations. Clinical examination of the Cameroonian family members failed to identify any symptoms previously reported in rare individuals carrying homozygous loss-of-function mutations in this gene. AP4E1 encodes the ε subunit of the heterotetrameric (ε-β4-μ4-σ4) AP-4 complex, involved in protein sorting at the trans-Golgi network. We found that the μ4 subunit of AP-4 interacts with NAGPA, an enzyme involved in the synthesis of the mannose 6-phosphate signal that targets acid hydrolases to the lysosome and the product of a gene previously associated with stuttering. These findings implicate deficits in intracellular trafficking in persistent stuttering.     

Light and electron microscopic immunocytochemical localization of two major proteins in garlic bulb

Garlic is known as a potent spice and a medicine with broad therapeutic properties ranging from antibacterial to anticancer, antidiabetic, and anticoagulant. Two major proteins of 40 KD and 14 KD constituting approximately 96% of total garlic proteins have been recently purified at our Institute. This immunocytochemical and ultrastructural study revealed that the 40 KD protein was localized in the parenchyma sheath cells (PSC) of garlic bulbs, whereas the 14 KD protein was present in the cortical cells (CC). Immunogold electron microscopy study indicated that the 40 KD protein was specifically localized in the globular granules of the cytoplasmic area of PSC. Each globular granule was amorphous and homogenous with membrane limiting its outermost layer. The yellowish color of PSC in freshly cut slices of garlic bulb suggested that PSC may have sulfur-containing compounds such as allicin, the primary contributor of the pungency and medicinal properties of garlic. Ellman's reagent test quantitatively revealed that there were 17.8 n moles sulfhydryl (SH)/ml of 40 KD garlic protein. Microtubule tubulin in mitotic figures from PHA-stimulated human short-term whole blood cultures reacted strongly with antitubulin antibody but reacted negatively with anti-40 KD garlic protein antibodies and therefore was not related to the 40 KD garlic protein immunocytochemically.     

Glycyrrhizic acid (GA), a triterpenoid saponin glycoside alleviates ultraviolet-B irradiation-induced photoaging in human dermal fibroblasts

Glycyrrhizic acid (GA), a triterpenoid saponin glycoside from the roots and rhizomes of licorice is used in traditional and modern medicine for the treatment of numerous medical conditions including skin diseases and beauty care product. In the present study, we investigated the effect of GA against ultraviolet B (UVB) irradiation-induced photoaging in human dermal fibroblasts (HDFs) and its possible mechanism of action. HDFs were subjected to photoaging by sub-toxic dose of UVB (10 mj/cm(2)) irradiation. Cell viability, matrix metalloproteinase 1 (MMP1), pro-collagen 1, cellular and nuclear morphology, cell cycle, intracellular reactive oxygen species (ROS), caspase 3 and hyaluronidase inhibition assays were performed. Western blotting was used to evaluate the expression of NF-kappa B (NF-κB) and cytochrome-C proteins. GA treatment significantly inhibited photoaging. It achieved this by reducing ROS, NF-κB, cytochrome c, caspase 3 levels and inhibiting hyaluronidase enzyme. The main mechanism seems to be, most likely by blocking MMP1 activation by modulating NF-κB signaling. These findings may be useful for development of natural and safe photoprotective agents against UVB irradiation.     

Correlation of serum oestrogen with salivary calcium in post-menopausal women with and without oral dryness feeling

doi: 10.1111/j.1741‐2358.2011.00580.x
Correlation of serum oestrogen with salivary calcium in post‐menopausal women with and without oral dryness feeling Objective: The aim of this study was to verify the association between salivary calcium and serum oestrogen levels with oral dryness in post‐menopausal women. Also, the correlation between these variables was evaluated. Methods: A case‐control study was carried out on 60 selected menopausal women with and without oral dryness feeling (30 as case and 30 as control) conducted in the Department of Oral Medicine and Radiology, Maharishi Markandeshar University, Mullana, India. Paraffin‐stimulated saliva samples were obtained by expectoration. Salivary calcium concentrations were assessed colorimetrically using Arsenazo III reaction. The serum oestrogen concentration was measured using ELISA. Statistical analysis of Student’s t‐test and Pearson correlation was used. Results: There was significant difference in mean values of both salivary calcium concentration and serum oestrogen between case and control groups. The result obtained also showed that an inverse correlation was found between salivary calcium concentration and serum oestrogen levels in both the groups and in total sample size. Conclusion: Oral dryness in post‐menopausal women is associated with high levels of salivary calcium and low levels of serum oestrogen. The concentrations of salivary calcium and serum oestrogen are inversely correlated in post‐menopausal women, regardless of the presence or absence of oral dryness.