Characterization of the axon initial segment of mice substantia nigra dopaminergic neurons

The axon initial segment (AIS) is the site of initiation of action potentials and influences action potential waveform, firing pattern, and rate. In view of the fundamental aspects of motor function and behavior that depend on the firing of substantia nigra pars compacta (SNc) dopaminergic neurons, we identified and characterized their AIS in the mouse. Immunostaining for tyrosine hydroxylase (TH), sodium channels (Na_v) and ankyrin‐G (Ank‐G) was used to visualize the AIS of dopaminergic neurons. Reconstructions of sampled AIS of dopaminergic neurons revealed variable lengths (12–60 μm) and diameters (0.2–0.8 μm), and an average of 50% reduction in diameter between their widest and thinnest parts. Ultrastructural analysis revealed submembranous localization of Ank‐G at nodes of Ranvier and AIS. Serial ultrathin section analysis and 3D reconstructions revealed that Ank‐G colocalized with TH only at the AIS. Few cases of synaptic innervation of the AIS of dopaminergic neurons were observed. mRNA in situ hybridization of brain‐specific Na_v subunits revealed the expression of Na_v1.2 by most SNc neurons and a small proportion expressing Na_v1.6. The presence of sodium channels, along with the submembranous location of Ank‐G is consistent with the role of AIS in action potential generation. Differences in the size of the AIS likely underlie differences in firing pattern, while the tapering diameter of AIS may define a trigger zone for action potentials. Finally, the conspicuous expression of Na_v1.2 by the majority of dopaminergic neurons may explain their high threshold for firing and their low discharge rate.     

Tumour biomechanical response to the vascular disrupting agent ZD6126 in vivo assessed by magnetic resonance elastography

Background:

Magnetic resonance elastography (MRE) is an emerging imaging technique that affords non-invasive quantitative assessment and visualization of tissue mechanical properties in vivo.

Methods:

In this study, MRE was used to quantify (kPa) the absolute value of the complex shear modulus |G*|, elasticity G_d and viscosity G_l of SW620 human colorectal cancer xenografts before and 24 h after treatment with either 200 mg kg⁻¹ of the vascular disrupting agent ZD6126 (N-acetylcolchinol-O-phosphate) or vehicle control, and the data were compared with changes in water diffusivity measured by diffusion-weighted magnetic resonance imaging.

Results:

A heterogeneous distribution of |G*|, G_d and G_l was observed pre-treatment with an intertumoral coefficient of variation of 13% for |G*|. There were no significant changes in the vehicle-treated cohort. In contrast, ZD6126 induced a significant decrease in the tumour-averaged |G*| (P<0.01), G_d (P<0.01) and G_l (P<0.05), and this was associated with histologically confirmed central necrosis. This reduction in tumour viscoelasticity occurred at a time when no significant change in tumour apparent diffusion coefficient (ADC) was observed.

Conclusions:

These data demonstrate that MRE can provide early imaging biomarkers for treatment-induced tumour necrosis.     

Activation of toll-like receptor 4 is necessary for trauma hemorrhagic shock-induced gut injury and polymorphonuclear neutrophil priming

Interactions of toll-like receptors (TLRs) with nonmicrobial factors play a major role in the pathogenesis of early trauma-hemorrhagic shock (T/HS)-induced organ injury and inflammation. Thus, we tested the hypothesis that TLR4 mutant (TLR4 mut) mice would be more resistant to T/HS-induced gut injury and polymorphonuclear neutrophil (PMN) priming than their wild-type littermates and found that both were significantly reduced in the TLR4 mut mice. In addition, the in vivo and ex vivo PMN priming effect of T/HS intestinal lymph observed in the wild-type mice was abrogated in TLR4 mut mice as well the TRIF mut-deficient mice and partially attenuated in Myd88 mice, suggesting that TRIF activation played a more predominant role than MyD88 in T/HS lymph-induced PMN priming. Polymorphonuclear neutrophil depletion studies showed that T/HS lymph-induced acute lung injury was PMN dependent, because lung injury was totally abrogated in PMN-depleted animals. Because the lymph samples were sterile and devoid of endotoxin or bacterial DNA, we investigated whether the effects of T/HS lymph was related to endogenous nonmicrobial TLR4 ligands. High-mobility group box 1 protein 1, heat shock protein 70, heat shock protein 27, and hyaluronic acid all have been implicated in ischemia-reperfusion-induced tissue injury. None of these "danger" proteins appeared to be involved, because their levels were similar between the sham and shock lymph samples. In conclusion, TLR4 activation is important in T/HS-induced gut injury and in T/HS lymph-induced PMN priming and lung injury. However, the T/HS-associated effects of TLR4 on gut barrier dysfunction can be uncoupled from the T/HS lymph-associated effects of TLR4 on PMN priming.     

Molecular Detection and Characterization of Porcine Epidemic Diarrhea Virus and Porcine Aichivirus C Coinfection in México

Swine enteric viral infections are responsible for substantial economic losses in the pork industry worldwide. Porcine epidemic diarrhea (PEDV) is one of the main causative agents of diarrhea in lactating pigs, and reports of PEDV coinfection with other enteric viruses highlight the importance of viral interactions for disease presentation and outcomes. Using next-generation sequencing (NGS) and sequence analyses from samples taken from piglets with acute diarrhea, we explored the possible interactions between PEDV and other less reported pathogens. PEDV coinfection with porcine kobuvirus (PKV) was detected in 36.4% (27/74) of samples. Full genomes from porcine coronavirus and kobuvirus were obtained, as was a partial porcine sapovirus genome (PSaV). The phylogenetic results show the clustering of these strains corresponding to the geographical relationship. To our knowledge, this is the first full genome and isolation report for porcine kobuvirus in México, as well as the first phylogenetic analysis for porcine sapovirus in the country. The NGS approach provides a better perspective of circulating viruses and other pathogens in affected production units.     

Oral bisphosphonate-induced osteonecrosis: risk factors, prediction of risk using serum CTX testing, prevention, and treatment.

PURPOSE: To assess the risk and time course of oral bisphosphonate-induced osteonecrosis of the jaws. MATERIALS AND METHODS: Detailed data from 30 consecutive cases were compared with 116 cases due to intravenous aminobisphosphonates. RESULTS: Results in part noted a higher incidence related to alendronate (Fosamax; Merck, Whitehouse Station, NJ), a 94.7% predilection for the posterior mandible, and a 50% occurrence spontaneously, with the remaining 50% resulting from an oral surgical procedure, mostly tooth removals. Just over 53% of patients were taking their oral bisphosphonate for osteopenia, 33.3% for documented osteoporosis, and 13.4% for steroid-induced osteoporosis related to 4 or more years of prednisone therapy for an autoimmune condition. There was a direct exponential relationship between the size of the exposed bone and the duration of oral bisphosphonate use. There was also a direct correlation between reports of pain and clinical evidence of infection. The morning fasting serum C-terminal telopeptide (CTX) test results were observed to correlate to the duration of oral bisphosphonate use and could indicate a recovery of bone remodeling with increased values if the oral bisphosphonate was discontinued. A stratification of relative risk was seen as CTX values less than 100 pg/mL representing high risk, CTX values between 100 pg/mL and 150 pg/mL representing moderate risk, and CTX values above 150 pg/mL representing minimal risk. The CTX values were noted to increase between 25.9 pg/mL to 26.4 pg/mL for each month of a drug holiday indicating a recovery of bone remodeling and a guideline as to when oral surgical procedures can be accomplished with the least risk. In addition, drug holidays associated with CTX values rising above the 150 pg/mL threshold were observed to correlate to either spontaneous bone healing or a complete healing response after an office-based debridement procedure. CONCLUSIONS: Oral bisphosphonate-induced osteonecrosis is a rare but real entity that is less frequent, less severe, more predictable, and more responsive to treatment than intravenous bisphosphonate-induced osteonecrosis. The morning fasting serum C-terminal telopeptide bone suppression marker is a useful tool for the clinician to assess risks and guide treatment decisions.     

Serologic survey of domestic animals for zoonotic arbovirus infections in the Lacandón Forest region of Chiapas,Mexico

A serologic survey in domestic animals (birds and mammals) was conducted in four communities located in the Lacandón Forest region of northeastern Chiapas, Mexico, during June 29 to July 1, 2001, with the objective to identify zoonotic arboviruses circulating in this area. We collected 202 serum samples from healthy domestic chickens, geese, ducks, turkeys, horses and cattle. The samples were tested by plaque-reduction neutralization test for antibodies to selected mosquito-borne flaviviruses (family Flaviviridae), including St. Louis encephalitis (SLE), Rocio (ROC), Ilheus (ILH), Bussuquara (BSQ), and West Nile (WN) viruses, and selected alphaviruses (family Togaviridae), including Western equine encephalitis (WEE), Eastern equine encephalomyelitis (EEE), and Venezuelan equine encephalitis (VEE) viruses. Neutralizing antibodies to SLE virus were detected in two (8%) of 26 turkeys, 15 (23%) of 66 cattle, and three (60%) of five horses. Antibodies to VEE virus were detected in 29 (45%) of 65 cattle. Because some of these animals were as young as 2 months old, we demonstrated recent activity of these two viruses. Sub-typing of the VEE antibody responses indicated that the etiologic agents of these infections belonged to the IE variety of VEE, which has been reported from other regions of Chiapas. WN virus-neutralizing antibodies were detected in a single cattle specimen (PRNT(90) = 1:80) that also circulated SLE virus-neutralizing antibodies (PRNT(90) = 1:20), suggesting that WN virus may have been introduced into the region. We also detected weak neutralizing activity to BSQ virus in four cattle and a chicken specimen, suggesting the presence of this or a closely related virus in Mexico. There was no evidence for transmission of the other viruses (ROC, ILH, EEE, WEE) in the study area.     

Association of N‐cadherin levels and downstream effectors of Rho GTPases with dendritic spine loss induced by chronic stress in rat hippocampal neurons

Chronic stress promotes cognitive impairment and dendritic spine loss in hippocampal neurons. In this animal model of depression, spine loss probably involves a weakening of the interaction between pre‐ and postsynaptic cell adhesion molecules, such as N‐cadherin, followed by disruption of the cytoskeleton. N‐cadherin, in concert with catenin, stabilizes the cytoskeleton through Rho‐family GTPases. Via their effector LIM kinase (LIMK), RhoA and ras‐related C3 botulinum toxin substrate 1 (RAC) GTPases phosphorylate and inhibit cofilin, an actin‐depolymerizing molecule, favoring spine growth. Additionally, RhoA, through Rho kinase (ROCK), inactivates myosin phosphatase through phosphorylation of the myosin‐binding subunit (MYPT1), producing actomyosin contraction and probable spine loss. Some micro‐RNAs negatively control the translation of specific mRNAs involved in Rho GTPase signaling. For example, miR‐138 indirectly activates RhoA, and miR‐134 reduces LIMK1 levels, resulting in spine shrinkage; in contrast, miR‐132 activates RAC1, promoting spine formation. We evaluated whether N‐cadherin/β‐catenin and Rho signaling is sensitive to chronic restraint stress. Stressed rats exhibit anhedonia, impaired associative learning, and immobility in the forced swim test and reduction in N‐cadherin levels but not β‐catenin in the hippocampus. We observed a reduction in spine number in the apical dendrites of CA1 pyramidal neurons, with no effect on the levels of miR‐132 or miR‐134. Although the stress did not modify the RAC–LIMK–cofilin signaling pathway, we observed increased phospho‐MYPT1 levels, probably mediated by RhoA–ROCK activation. Furthermore, chronic stress raises the levels of miR‐138 in accordance with the observed activation of the RhoA–ROCK pathway. Our findings suggest that a dysregulation of RhoA–ROCK activity by chronic stress could potentially underlie spine loss in hippocampal neurons. © 2015 Wiley Periodicals, Inc.