Establishment of pseudorabies virus latency and reactivation model in mice dorsal root ganglia culture.

Pseudorabies virus (PRV) belongs to the alpha herpesvirus family and is responsible for Aujeszky's disease in pigs. Similar to other alpha herpesviruses, PRV establishes a lifelong latent infection in trigeminal ganglion. These latently infected pigs serve as a reservoir for recurrent infections when reactivation is triggered, making the eradication of PRV a challenging task. However, the molecular mechanism underlying PRV latency and reactivation in neurons is still poorly understood due to limitations in the in vitro model. To establish a pseudorabies virus latency and reactivation model in primary neuron cultures, we isolated dorsal root ganglion (DRG) from newborn Kunming mice using a method named epineurium-pulling for DRG collection (EPDC) and cultured primary neurons in vitro. A dual-colour recombinant PRV BAC mRuby-VP16 was constructed and 0.5 multiplicity of infection (MOI) was found as an appropriate dose in the presence of aciclovir to establish latency. Reactivation was induced using UV-inactivated herpesviruses or a series of chemical inhibitors. Interestingly, we found that not only UV-PRV, but also UV-HSV-1 and UV-BHoV-5 were able to induce rapid PRV reactivation. The efficiency of reactivation for LY294002, forskolin, etoposide, dexamethasone, and acetylcholine was found to be dependent on their concentration. In conclusion, we developed a valuable model of PRV latency and reactivation, which provides a basis for future mechanism research.

Long-Term Outcomes of Total Hip Arthroplasty With Transverse Subtrochanteric Shortening Osteotomy and Modular Stem in Crowe IV Developmental Dysplasia.

BACKGROUND: The reconstruction of high dislocation related to developmental dysplasia of the hip (DDH) remains challenging for joint surgeons. The aim of this study is to evaluate the rate of union, the revision rate, functional scores, and complications in patients with Crowe IV DDH treated with total hip arthroplasty, transverse subtrochanteric shortening osteotomy, and modular stem in an average 10-year follow-up. METHODS: Twenty-eight patients (33 hips) with Crowe IV DDH who were operated on between 2008 and 2013 were followed. All patients underwent uncemented total hip arthroplasty with transverse subtrochanteric shortening osteotomy and anatomical acetabular cup implantation. The mean age was 36.6 years, and the mean follow-up period was 121 months. Clinical and radiological outcomes were evaluated. RESULTS: The mean Harris Hip Score significantly improved from 47.0 preoperatively to 89.6 postoperatively. The mean limb length discrepancy was significantly reduced from 3.8 to 0.8 cm. The mean osteotomy union time was 6.8 months. At the mean follow-up of 121 months, there were 3 cases of postoperative dislocation, 2 cases of intraoperative fracture, and 1 case of posterior tibial venous thrombosis. No revision occurred, and no signs of component loosening or migration were observed at the last follow-up. CONCLUSION: Crowe IV DDH patients treated with transverse subtrochanteric shortening osteotomy, modular stem, and anatomic acetabular component insertion can have satisfactory and reliable 10-year clinical outcomes.

Typical gene expression profile of pseudorabies virus reactivation from latency in swine trigeminal ganglion.

Pseudorabies virus (PRV) establishes a lifelong latent infection in swine trigeminal ganglion (TG) following acute infection. Increased corticosteroid levels, due to stress, increases the incidence of reactivation from latency. Muscle injection combined with intravenous deliver of the synthetic corticosteroid dexamethasone (DEX) consistently induces reactivation from latency in pigs. In this study, PRV-free piglets were infected with PRV. Viral shedding in nasal and ocular swabs demonstrated that PRV infection entered the latent period. The anti-PRV antibody was detected by enzyme-linked immunosorbent assay and the serum neutralization test, which suggested that the PRV could establish latent infection in the presence of humoral immunity. Immunohistochemistry and viral genome detection of TG neurons suggested that PRV was reactivated from latency. Viral gene expressions of IE180, EP0, VP16, and LLT-intron were readily detected at 3-h post-DEX treatment, but gB, a γ1 gene, was not detectable. The differentially expressed phosphorylated proteins of TG neurons were analyzed by ITRAQ coupled with LC-MS/MS, and p-EIF2S2 differentially expression was confirmed by western blot assay. Taken together, our study provides the evidence that typical gene expression in PRV reactivation from latency in TG is disordered compared with known lytic infection in epithelial cells.

Pseudorabies virus UL13 primes inflammatory response through downregulating heat shock factor 1.

Pseudorabies virus is a swine alpha-herpesvirus. We demonstrated that alpha-herpesvirus infection downregulates HSF1, a master transcription factor in the heat shock response. The serine/threonine protein kinase activity of late viral protein UL13 is indispensable for HSF1 depletion and phosphorylation, and UL13 does not degrade HSF1 posttranslationally but inhibits the HSF1 mRNA level. Importantly, UL13 increased HSF1 activity even though it reduced HSF1 mRNA. Furthermore, viral replication markedly decreased in the HSF1 knockout cell line or in the presence of an HSF1-specific inhibitor. Interestingly, HSF1 knockout accelerated the activation of NF-κB and p38MAPK. The K96 loci of UL13 are important to induce high levels of IL-6, TNF-α, and IL-ß cytokines while playing a crucial role in promoting mild interstitial pneumonia, liver necrosis, and severe inflammatory cell infiltration in the footpad. Thus, UL13 steers the heat shock response to promote viral replication and the inflammatory response. IMPORTANCE: PRV is a ubiquitous pathogen that infects a variety of mammals, such as pigs, ruminants, carnivores, and rodents as well as human beings, causing enormous economic losses in the swine industry. Here, we employed PRV as a model to determine the relationship between α-herpesvirus and the inflammatory response. Overall, our findings indicated that PRV infection inhibits the level of HSF1 mRNA via the serine/threonine protein kinase activity of UL13. Additionally, we discovered that HSF1 was involved in NF-κB activation upon PRV infection. PRV UL13 orchestrates the level of HSF1 mRNA, HSF1 protein phosphorylation, and priming of the inflammatory response. Our study reveals a novel mechanism employed by UL13 serine/threonine protein kinase activity to promote the inflammatory response, providing novel clues for therapy against alpha-herpesvirus infection.

Hsp90 is involved in pseudorabies virus virion assembly via stabilizing major capsid protein VP5.

Many viruses utilize molecular chaperone heat shock protein 90 (Hsp90) for protein folding and stabilization, however, the role of Hsp90 in herpesvirus lifecycle is obscure. Here, we provide evidence that Hsp90 participates in pseudorabies virus (PRV) replication. Viral growth kinetics assays show that Hsp90 inhibitor geldanamycin (GA) abrogates PRV replication at the post-penetration step. Transmission electron microscopy demonstrates that dysfunction of Hsp90 diminishes the quantity of PRV nucleocapsids. Overexpression and knockdown of Hsp90 suggest that de novo Hsp90 is involved in PRV replication. Mechanismly, dysfunction of Hsp90 inhibits PRV major capsid protein VP5 expression. Co-immunoprecipitation and indirect immunofluorescence assays indicate that Hsp90 interacts with VP5. Interestingly, Hsp70, a collaborator of Hsp90, also interacts with VP5, but doesn't affect PRV growth. Finally, inhibition of Hsp90 results in PRV VP5 degradation in a proteasome-dependent manner. Collectively, our data suggest that Hsp90 contributes to PRV virion assembly and replication via stabilization of VP5.

How much space of the spinal canal should be restored by hoisting the vertebrae-OPLL complex for sufficient decompression in anterior controllable antedisplacement and fusion? A multicenter clinical radiological study.

BACKGROUND CONTEXT: Anterior controllable antedisplacement and fusion (ACAF) is a novel surgical technique for the treatment of ossification of the posterior longitudinal ligament (OPLL). Its prognostic factors for decompression have not been well studied. Additionally, no detailed radiological standard has been set for hoisting the vertebrae-OPLL complex (VOC) in ACAF. PURPOSE: To identify the possible prognostic factors for decompression outcomes after ACAF for cervical OPLL, to determine the critical value of radiological parameters for predicting good outcomes, and to establish a radiological standard for hoisting the VOC in ACAF. STUDY DESIGN: This was a retrospective multicenter study. PATIENT SAMPLE: A total of 121 consecutive patients with OPLL who underwent ACAF at a point between January 2017 and June 2018 at any one of seven facilities and were monitored for at least 1 year afterward were enrolled in a multicenter study. OUTCOME MEASURES: Japanese Orthopedic Association (JOA) scores, recovery rate (RR) of neurologic function, and surgical complications were used to determine the effectiveness of ACAF. METHODS: Patients were divided into two groups according to their RR for neurologic function. Patients with an RR of ≥50% and an RR of <50% were designated as having good and poor decompression outcomes, respectively. The relationship between various possible prognostic factors and decompression outcomes was assessed by univariate and multivariate analysis. The receiver operating characteristic curve was used to determine the optimal cutoff value of the radiological parameters for prediction of good decompression outcomes. Next, the patients were redivided into three groups according to the cutoff value of the selected radiological parameter (postoperative anteroposterior canal diameter [APD] ratio). Patients with postoperative APD ratios of ≤80.7%, 80.7%-100%, and ≥100% were defined as members of the incomplete, optimal, and excessive antedisplacement groups, respectively. Differences in decompression outcomes among the three groups were compared to verify the reliability of the postoperative APD ratio and assess the necessity of excessive antedisplacement. RESULTS: Multivariate logistic regression analysis showed that patients' age at surgery (odds ratio [OR]=1.18; 95% confidence interval [CI]=1.08-1.29; p<.01) and postoperative APD ratio (OR=0.83; 95% CI=0.77-0.90; p<.01) were independently associated with decompression outcomes. The optimal cutoff point of the postoperative APD ratio was calculated at 80.7%, with 86.2% sensitivity and 73.5% specificity. There were no significant differences in the postoperative JOA scores and RRs between the excessive antedisplacement group and optimal antedisplacement group (p>.05). However, a lower incidence of cerebrospinal fluid leakage and screw slippage was observed in the optimal antedisplacement group (p<.05). CONCLUSIONS: Patients' age at surgery and their postoperative APD ratio are the two prognostic factors of decompression outcomes after ACAF. The postoperative APD ratio is also the most accurate radiological parameter for predicting good outcomes. Our findings suggest that it is essential for neurologic recovery to restore the spinal canal to more than 80.7% of its original size (postoperative APD ratio >80.7%), and restoration to less than its original size (postoperative APD ratio <100%) will help reduce the incidence of surgical complications. This may serve as a valuable reference for establishment of a radiological standard for hoisting the VOC in ACAF.

[Study on the influence of mineralizer on the structures and spectral properties of calcium aluminates].

The present paper investigated the effect of mineralizer on the structure and properties of calcium aluminates formation. Calcium aluminates powder was synthesized under high temperature calcination by mixing bauxite, limestone and a certain amount of mineralizer. The product structure, compositional information and spectral properties were carefully characterized by XRD, IR and DTA-TG, and the mineralization mechanism of mineralizer was studied during the process of calcium aluminates preparation. The results showed that calcium aluminates powder could be obtained under lower temperature calcination after adding mineralizer to the raw materials. The main products of the reaction were CaAl10 O18 and CaAl2 Si2 O8 without mineralizer, however, the main products of the reaction were CaAl3 BO7 and Ca3 Al10 O18 with mineralizer, in which Al2 O3s could be extracted easily, while CaAl2 Si2 O8 was reduced greatly in which Al2 O3 could not be extracted easily. At the same time, it is easy for calcspar to decompose after adding mineralizer. It is favorable to Al-Si bond break and Al stripping from bauxite. These facts could improve the extraction rate of Al2 O3 from raw materials. Also, in the case of adding mineralizer to the raw mixes, the crystal structure and composition are changed, which is beneficial to reducing calcination temperature.

Effect of ZBD-2 on chronic pain, depressive-like behaviors, and recovery of motor function following spinal cord injury in mice.

In addition to debilitating sensory and motor deficits, patients with spinal cord injury (SCI) may experience chronic hyperpathic pain (SCI-pain). Recent studies have revealed that translocator protein (TSPO) is involved in repairing neural cells as well as reducing anxiety and depression. However, the role of TSPO in SCI-pain and pain-induced depression remains unknown. The present study aimed to determine the effects of a new TSPO ligand, ZBD-2, on SCI-pain and consequent pain-induced depressive-like behaviors in mice. Treatment with ZBD-2 at either dose significantly attenuated the symptoms of chronic SCI-pain and pain-induced depressive-like behaviors. ZBD-2 reversed SCI-induced elevation of serum corticosterone levels, an index of hyper-activation of the hypothalamic-pituitary-adrenal (HPA) axis. Additionally, administration of ZBD-2 inhibited decreases in the expression of synaptic plasticity-related signaling proteins, including brain-derived neurotrophic factor (BDNF) and cyclic AMP-responsive element binding protein (CREB). Moreover, ZBD-2 administration reversed chronic, SCI-induced gliocyte activation at the lesion site. Therefore, ZBD-2 may improve chronic SCI-pain and pain-induced depressive-like behaviors via suppression of gliocyte activation and restoration of the synaptic plasticity-related signaling systems.

A dual-heterodyne laser interferometer for simultaneous measurement of linear and angular displacements.

Picometer laser interferometry is an essential tool for ultra-precision measurements in frontier scientific research and advanced manufacturing. In this paper, we present a dual-heterodyne laser interferometer for simultaneously measuring linear and angular displacements with resolutions of picometer and nanoradian, respectively. The phase measurement method is based on cross-correlation analysis and realized by a PXI-bus data acquisition system. By implementing a dual-heterodyne interferometer with a highly symmetric optical configuration, low frequency noises caused by the environmental fluctuations can be suppressed to very low levels via common-mode noise rejection. Experimental results for the dual-heterodyne interferometer configuration presented demonstrate that the noise levels of the linear and angular displacement measurements are approximately 1 pm/Hz(1/2) and 0.5 nrad/Hz(1/2) at 1 Hz.