Weekly administration of teriparatide has been shown to reduce the risk of vertebral and non-vertebral fractures in patients with osteoporosis at higher fracture risk in Japan. However, its efficacy for hip fracture has not been established. To gain insight into the effect of weekly teriparatide on the hip, hip structural analysis (HSA) based on dual-energy X-ray absorptiometry (DXA) was performed using the data of 209 postmenopausal osteoporotic women who had participated in the original randomized, multicenter, double-blind, placebo-controlled trial assessing the effects of once-weekly 56.5 ?g teriparatide for 72 weeks. The DXA scans, obtained at baseline, 48 weeks and 72 weeks, were analyzed to extract bone mineral density (BMD) and cross-sectional geometrical indices at the narrowest point on the neck (NN), the intertrochanteric region (IT), and the proximal shaft. Compared with placebo after 72 weeks, the teriparatide group showed significantly higher BMD, average cortical thickness, bone cross-sectional area, and section modulus, and lower buckling ratio at both the NN and IT regions. No significant expansion of periosteal diameter was observed at these regions. There were no significant differences in BMD and HSA indices at the shaft region. The results indicate that overall structural strength in the proximal femur increased compared to placebo, suggesting that once-weekly teriparatide effectively reverses changes in hip geometry and strength with aging.
The axon initial segment (AIS) is a structurally and molecularly unique neuronal compartment of the proximal axon that functions as both a physiological and physical bridge between the somatodendritic and axonal domains. The AIS has two main functions: to initiate action potentials and to maintain neuronal polarity. The cytoskeletal scaffold ankyrinG is responsible for these functions and clusters ion channels at the AIS. Recent studies reveal how the AIS forms and remarkable diversity in its structure, function, and composition that may be modulated by neuronal activity and posttranslational modifications of AIS proteins. Furthermore, AIS proteins have been implicated in a variety of human diseases. Here, we discuss these findings and what they teach us about the dynamic AIS.
Lewis X (Le(X), Gal?1-4(Fuc?1-3)GlcNAc) is a carbohydrate epitope that is present at the nonreducing terminus of sugar chains of glycoproteins and glycolipids, and is abundantly expressed in several stem cell populations. Le(X) antigen can be used in conjunction with fluorescence-activated cell sorting to isolate neurosphere-forming neural stem cells (NSCs) from embryonic mouse brains. However, its function in the maintenance and differentiation of stem cells remains largely unknown. In this study, we examined mice deficient for fucosyltransferase 9 (Fut9), which is thought to synthesize most, if not all, of the Le(X) moieties in the brain. We found that the number of NSCs was increased in the brain of Fut9(-/-) embryos, suggesting that Fut9-synthesized Le(X) is dispensable for the maintenance of NSCs. Another ?1,3-fucosyltransferase gene, fucosyltransferase 10 (Fut10), is expressed in the ventricular zone of the embryonic brain. Overexpression of Fut10 enhanced the self-renewal of NSCs. Conversely, suppression of Fut10 expression induced the differentiation of NSCs and embryonic stem cells. In addition, knockdown of Fut10 expression in the cortical ventricular zone of the embryonic brain by in utero electroporation of Fut10-miRNAs impaired the radial migration of neural precursor cells. Our data suggest that Fut10 is involved in a unique ?1,3-fucosyltransferase activity with stringent substrate specificity, and that this activity is required to maintain stem cells in an undifferentiated state.
Histo-blood group antigens (HBGAs) have been suggested to be receptors or coreceptors for human noroviruses (HuNoVs) expressed on the intestinal epithelium. We isolated an enteric bacterium strain (SENG-6), closely related to Enterobacter cloacae, bearing HBGA-like substances from a fecal sample of a healthy individual by using a biopanning technique with anti-HBGA antibodies. The binding capacities of four genotypes of norovirus-like particles (NoVLPs) to Enterobacter sp. SENG-6 cells were confirmed by enzyme-linked immunosorbent assay (ELISA). Transmission electron microscopy demonstrated that NoVLPs bound mainly to extracellular polymeric substances (EPS) of Enterobacter sp. SENG-6, where the HBGA-like substances were localized. EPS that contained HBGA-like substances extracted from Enterobacter sp. SENG-6 was shown by enzyme-linked immunosorbent assay (ELISA) to be capable of binding to NoVLPs of a GI.1 wild-type strain (8fIIa) and a GII.6 strain that can recognize A antigen but not to an NoVLP GI.1 mutant strain (W375A) that loses the ability to bind to A antigen. Enzymatic cleavage of terminal N-acetyl-galactosamine residues in the bacterial EPS weakened bacterial EPS binding to the GI.1 wild-type strain (8fIIa). These results indicate that A-like substances in the bacterial EPS play a key role in binding to NoVLPs. Since the specific binding of HuNoVs to HBGA-positive enteric bacteria is likely to affect the transmission and infection processes of HuNoVs in their hosts and in the environment, further studies of human enteric bacteria and their binding capacity to HuNoVs will provide a new scientific platform for understanding interactions between two types of microbes that were previously regarded as biologically unrelated.
Specific growth rates and hydrocarbon contents of Botryococcus braunii strain Showa were measured under a wide range of CO2, salinity, temperature, and irradiance conditions. The bubbling CO2 concentration of 0.2-5% and no addition of salinity were favorable conditions for growth. The strain cannot grow at 5°C and above 35°C under any irradiance levels. Maximum specific growth rate of 0.5 day(-1) (doubling time of 1.4 days), the highest value reported for B. braunii in the past studies, was observed at 30°C and 850 ?mol photons m(-2) s(-1). Since hydrocarbon productivity, shown as the product of hydrocarbon content and specific growth rate, increased with the increasing specific growth rate, we conclude that more efficient hydrocarbon production by the mass culture of strain Showa can be achieved by maintaining higher specific growth rate based on the culture conditions presented in this study.
Semaphorin3A (Sema3A) is a repulsive guidance molecule for axons, which acts by inducing growth cone collapse through phosphorylation of CRMP2 (collapsin response mediator protein 2). Here, we show a role for CRMP2 oxidation and thioredoxin (TRX) in the regulation of CRMP2 phosphorylation and growth cone collapse. Sema3A stimulation generated hydrogen peroxide (H2O2) through MICAL (molecule interacting with CasL) and oxidized CRMP2, enabling it to form a disulfide-linked homodimer through cysteine-504. Oxidized CRMP2 then formed a transient disulfide-linked complex with TRX, which stimulated CRMP2 phosphorylation by glycogen synthase kinase-3, leading to growth cone collapse. We also reconstituted oxidation-dependent phosphorylation of CRMP2 in vitro, using a limited set of purified proteins. Our results not only clarify the importance of H2O2 and CRMP2 oxidation in Sema3A-induced growth cone collapse but also indicate an unappreciated role for TRX in linking CRMP2 oxidation to phosphorylation.
In addition to its regulatory effect on bone mass, calcitonin has been shown to relieve pain and alleviate peripheral circulatory disturbance in patients with Raynauds syndrome and complex regional pain syndrome. In the present study, we investigated whether calcitonin ameliorates diminished blood flow and enhanced arterial contraction in response to noradrenaline in chronic constriction injury (CCI) of the sciatic nerve in rats. Following surgically induced CCI, laser Doppler flowmetry studies showed a significant decrease in plantar skin blood flow of the ipsilateral hind paw compared to the contralateral side. A subcutaneous bolus injection of elcatonin (20 U/kg), a synthetic derivative of eel calcitonin, significantly improved decreased skin blood flow in the ipsilateral side. In vitro analysis of plantar arteries isolated from the ipsilateral hind paw 7-13 days after the CCI procedure showed higher sensitivity to noradrenaline than the plantar arteries from the contralateral side. Elcatonin (0.1-10 nm) significantly reduced noradrenaline-induced contraction in the arteries of the ipsilateral side, whereas it had little effect on those of the contralateral side. These results suggest that calcitonin selectively ameliorates enhanced arterial contractility in CCI neuropathic rats, thus leading to its alleviating effect on peripheral circulatory disturbance.
The correct balance between excitation and inhibition is crucial for brain function and disrupted in several pathological conditions. Excitatory neuronal circuits in the primary somatosensory cortex (S1) are modulated by local inhibitory neurons with the balance of this excitatory and inhibitory activity important for function. The activity of excitatory layer 2/3 neurons (L2/3) in the S1 cortex is increased in chronic pain, but it is not known how the local interneurons, nor the balance between excitation and inhibition, may change in chronic pain. Using in vivo two-photon calcium imaging and electrophysiology, we report here that the response of L2/3 local inhibitory neurons to both sensory stimulation and to layer 4 electrical stimulation increases in inflammatory chronic pain. Local application into L2/3 of a GABA(A) receptor blocker further enhanced the activity of S1 excitatory neurons and reduced pain thresholds, whereas local application of the GABA(A) receptor modulators (muscimol and diazepam) transiently alleviated the allodynia. This illustrates the importance of the local inhibitory pathways in chronic pain sensation. A reduction in the expression and function of the potassium-chloride cotransporter 2 occurred during chronic pain, which reduces the efficacy of the inhibitory inputs to L2/3 excitatory neurons. In summary, both excitatory and inhibitory neuronal activities in the S1 are enhanced in the chronic pain model, but the increased inhibition is insufficient to completely counterbalance the increased excitation and alleviate the symptoms of chronic pain.
The polypeptide hormone calcitonin is clinically well known for its ability to relieve neuropathic pain such as spinal canal stenosis, diabetic neuropathy and complex regional pain syndrome. Mechanisms for its analgesic effect, however, remain unclear. Here we investigated the mechanism of anti-hyperalgesic action of calcitonin in a neuropathic pain model in rats.
We have established 3T3-L1 cells possessing a secretory Gaussia luciferase (GLuc) gene under the control of nuclear factor-kappa B (NF-?B) response element. The 3T3-L1 cells named 3T3-L1-NF-?B-RE-GLuc could differentiate into adipocyte as comparably as parental 3T3-L1 cells. Inflammatory cytokines such as tumor necrosis factor (TNF)-? and interleukin (IL)-1? induced GLuc secretion of 3T3-L1-NF-?B-RE-GLuc adipocytes in a concentration- and time-dependent manner. GLuc secretion of 3T3-L1-NF-?B-RE-GLuc adipocytes was also induced when cultured with RAW264.7 macrophages and was dramatically enhanced by lipopolysaccharide (LPS)-activated macrophages. An NF-?B activation inhibitor BAY-11-7085 and an antioxidant N-acetyl cysteine significantly suppressed GLuc secretion induced by macrophages. Finally, we found that rosemary-derived carnosic acid strongly suppressed GLuc secretion induced by macrophages and on the contrary up-regulated adiponectin secretion. Collectively, by using 3T3-L1-NF-?B-RE-GLuc adipocytes, inflammation status can be monitored in real time and inflammation-attenuating compounds can be screened more conveniently.
N-linked glycans harbored on glycoproteins profoundly affect the character of proteins by altering their structure or capacity to bind to other molecules. Specific knowledge of the role of N-glycans in these changes is limited due to difficulties in identifying precise carbohydrate structures on a given glycoprotein, which arises from the large amounts of glycoprotein required for N-glycan structural determination. Here, we refined a simple method to purify and detect trace amounts of N-glycans. During the N-glycan purification step, most contaminants were removed by two kinds of columns: a graphite carbon column and a cellulose column. N-Glycans were identified with a three-dimensional high-performance liquid chromatography (HPLC) system. Using our method, a global analysis of N-glycans from human muscle biopsy samples and mouse brain sections was possible. By combining sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with our method, we refined analytical procedures for N-glycans from SDS-PAGE gels using hydrazinolysis to achieve a high N-glycan recovery rate. N-Glycans on as little as 1 ?g of the target protein transferrin or immunoglobulin G (IgG) were easily detected. These methods allowed us to efficiently determine glycoprotein N-glycans at picomole (pmol) levels.
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