Alpha-1 Antitrypsin-Induced Endoplasmic Reticulum Stress Promotes Invasion by Extravillous Trophoblasts.

Alpha-1 antitrypsin (A1AT) is a glycoprotein that has been shown to protect tissues from proteolytic damage under various inflammatory conditions. Several studies show that A1AT may be associated with pre-eclampsia. However, the role of A1AT expression in placental physiology is not fully understood. In the present study, we aim to characterize the expression and function of placental A1AT. A1AT knockdown is found to reduce the expression of the serine protease HTRA1 in a trophoblast cell line. In addition, A1AT overexpression (A1AT-OE) increases the expression of HTRA1, IL6, CXCL8, and several markers of endoplasmic reticulum (ER) stress. Treatment with tunicamycin or thapsigargin, which induces ER stress, increases HTRA1 expression. Furthermore, immunohistochemistry reveals that HTRA1 is expressed in trophoblasts and the endometrial decidual cells of human placentas. An invasion assay shows that A1AT and HTRA1 stimulate cell invasion, but treatment with the ER stress inhibitors reduces the expression of HTRA1 and ER stress markers and prevents cell invasion in A1AT-OE trophoblasts. These results suggest that endogenous A1AT regulates inflammatory cytokine expression and HTRA1-induced trophoblast invasion via the induction of ER stress. It is concluded that an imbalance in the functional link between A1AT and ER stress at the maternal-fetal interface might cause abnormal placental development.

Cytotoxin-mediated silk gland organ dysfunction diverts resources to enhance silkworm fecundity by potentiating nutrient-sensing IIS/TOR pathways.

Energy reserves, primarily stored in the insect's fat body, are essential for physiological processes such as reproduction and cocoon formation. However, whether these processes are mutually constraining is unknown. Here, we showed that cocoon-free silkworms accumulate amino acid constituents of silk proteins in the hemolymph and maintain lipid and sugar reserves in the pupal fat body by repressing the expression of sericin and fibroin genes in the middle and posterior silk glands, respectively, via butterfly pierisin-1A catalytic domain expression. This, in turn, upregulates insulin/insulin-like signaling and target of rapamycin (IIS/TOR) signaling, which enhances vitellogenesis and accelerates ovarian development, thus contributing to increased fecundity. The impacts of semi-starvation on fecundity and egg hatchability were also less pronounced in cocoon-free silkworms compared with wildtype silkworms. These data uncover the resource allocation trade-off between cocoon formation and fecundity and demonstrate that nutritional signaling plays a role in regulating silkworm reproduction.

Enhancement of bone regeneration by coadministration of angiogenic and osteogenic factors using messenger RNA.

BACKGROUND: Bone defects remain a challenge today. In addition to osteogenic activation, the crucial role of angiogenesis has also gained attention. In particular, vascular endothelial growth factor (VEGF) is likely to play a significant role in bone regeneration, not only to restore blood supply but also to be directly involved in the osteogenic differentiation of mesenchymal stem cells. In this study, to produce additive angiogenic-osteogenic effects in the process of bone regeneration, VEGF and Runt-related transcription factor 2 (Runx2), an essential transcription factor for osteogenic differentiation, were coadministered with messenger RNAs (mRNAs) to bone defects in the rat mandible. METHODS: The mRNAs encoding VEGF or Runx2 were prepared via in vitro transcription (IVT). Osteogenic differentiation after mRNA transfection was evaluated using primary osteoblast-like cells, followed by an evaluation of the gene expression levels of osteogenic markers. The mRNAs were then administered to a bone defect prepared in the rat mandible using our original cationic polymer-based carrier, the polyplex nanomicelle. The bone regeneration was evaluated by micro-computerized tomography (µCT) imaging, and histologic analyses. RESULTS: Osteogenic markers such as osteocalcin (Ocn) and osteopontin (Opn) were significantly upregulated after mRNA transfection. VEGF mRNA was revealed to have a distinct osteoblastic function similar to that of Runx2 mRNA, and the combined use of the two mRNAs resulted in further upregulation of the markers. After in vivo administration into the bone defect, the two mRNAs induced significant enhancement of bone regeneration with increased bone mineralization. Histological analyses using antibodies against the Cluster of Differentiation 31 protein (CD31), alkaline phosphatase (ALP), or OCN revealed that the mRNAs induced the upregulation of osteogenic markers in the defect, together with increased vessel formation, leading to rapid bone formation. CONCLUSIONS: These results demonstrate the feasibility of using mRNA medicines to introduce various therapeutic factors, including transcription factors, into target sites. This study provides valuable information for the development of mRNA therapeutics for tissue engineering.

[Development and neural basis of social behavior].

We have developed a multi variate analysis of social communication behavior which allows for discriminating the emotional state of an agent interacting with other agents (Bouquet method). Domestic chick or common marmoset was reared under socially isolated conditions, later tested for its development of communication behavior with peers by Bouquet, and compared with that of animals reared under grouping conditions. We found the existence of high sensitive period for social interaction, given less experience, developing less affiliated behavior. From chick model, a couple of neuronal difference was observed between two groups, amygdale core central cell size, MRI volumetric measure in mesolimbic area, and gene expression patter including brain type tryptophan hydroxylase in nucleus accumbens. The isolated chick behavior changed better to be socially affiliated by taking SSRI/SNRI or Ubiquinol (the reduced form of CoQ10) together with social interaction experience after the sensitive period. Finally, we could discriminate the behavior of Asperger syndrome children (n = 7) from that of the typically developed siblings (n = 6) during the clinical interview by applying Bouquet method.

Anti-Inflammatory Therapy for Temporomandibular Joint Osteoarthritis Using mRNA Medicine Encoding Interleukin-1 Receptor Antagonist.

Messenger RNA (mRNA) is an emerging drug modality for protein replacement therapy. As mRNA efficiently provides protein expression in post-mitotic cells without the risk of insertional mutagenesis, direct delivery of mRNA can be applied, not only as an alternative to gene therapy, but also for various common diseases such as osteoarthritis (OA). In this study, using an mRNA-encoding interleukin-1 receptor antagonist (IL-1Ra), we attempted anti-inflammatory therapy in a rat model of the temporomandibular joint (TMJ) OA, which causes long-lasting joint pain with chronic inflammation. For the intra-articular injection of mRNA, a polyplex nanomicelle, our original polymer-based carrier, was used to offer the advantage of excellent tissue penetration with few immunogenic responses. While the protein expression was transient, a single administration of IL-1Ra mRNA provided sustained pain relief and an inhibitory effect on OA progression for 4 weeks. The mRNA-loaded nanomicelles provided the encoded protein diffusely in the disc and articular cartilage without upregulation of the expression levels of the pro-inflammatory cytokines IL-6 and tumor necrosis factor-α (TNF-α). This proof-of-concept study demonstrates how anti-inflammatory proteins delivered by mRNA delivery using a polyplex nanomicelle could act to alleviate OA, stimulating the development of mRNA therapeutics.

Brain Dp140 alters glutamatergic transmission and social behaviour in the mdx52 mouse model of Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a muscle disorder caused by DMD mutations and is characterized by neurobehavioural comorbidities due to dystrophin deficiency in the brain. The lack of Dp140, a dystrophin short isoform, is clinically associated with intellectual disability and autism spectrum disorders (ASDs), but its postnatal functional role is not well understood. To investigate synaptic function in the presence or absence of brain Dp140, we utilized two DMD mouse models, mdx23 and mdx52 mice, in which Dp140 is preserved or lacking, respectively. ASD-like behaviours were observed in pups and 8-week-old mdx52 mice lacking Dp140. Paired-pulse ratio of excitatory postsynaptic currents, glutamatergic vesicle number in basolateral amygdala neurons, and glutamatergic transmission in medial prefrontal cortex-basolateral amygdala projections were significantly reduced in mdx52 mice compared to those in wild-type and mdx23 mice. ASD-like behaviour and electrophysiological findings in mdx52 mice were ameliorated by restoration of Dp140 following intra-cerebroventricular injection of antisense oligonucleotide drug-induced exon 53 skipping or intra-basolateral amygdala administration of Dp140 mRNA-based drug. Our results implicate Dp140 in ASD-like behaviour via altered glutamatergic transmission in the basolateral amygdala of mdx52 mice.

Remarkable birefringence in a TiO2-SiO2 composite film with an aligned mesoporous structure.

Mesoporous titania-silica composite films with highly aligned cylindrical pores are prepared by the sol-gel method using a substrate with structural anisotropy. The strong alignment effect of a rubbing-treated polyimide film on a substrate provides a narrow alignment distribution in the plane of the film regardless of the fast condensation rate of titania precursors. The collapse of the mesostructure upon the surfactant removal is effectively suppressed by the reinforcement of the pore walls with silica by exposing the as-deposited film to a vapor of a silicon alkoxide. The existence of a silica layer on the titania pore wall is proved from the distributions of Ti and Si estimated by the elemental analysis in high resolution electron microscopy. The obtained mesoporous titania-silica composite film exhibits a remarkable birefringence reflecting the highly anisotropic mesoporous structure and the high refractive index of titania that forms the pore wall. The Δn value estimated from the optical retardation and the film thickness is larger than 0.06, which cannot be achieved with the conventional mesoporous silica films with uniaxially aligned mesoporous structure even though the alignment of the pores in the films is perfect. These inorganic films with mesoscopic structural anisotropy will find many applications in the field of optics as phase plates with high thermal/chemical/mechanical stabilities.

Treatment of ischemic neuronal death by introducing brain-derived neurotrophic factor mRNA using polyplex nanomicelle.

Ischemic neuronal death causes serious lifelong neurological deficits; however, there is no proven effective treatment that can prevent neuronal death after the ischemia. We investigated the feasibility of mRNA therapeutics for preventing the neuronal death in a rat model of transient global ischemia (TGI). By intraventricular administration of mRNA encoding brain-derived neurotrophic factor (BDNF) using a polymer-based carrier, polyplex nanomicelle, the mRNA significantly increased the survival rate of hippocampal neurons after TGI, with a rapid rise of BDNF in the hippocampus. Interestingly, mRNA administration on Day 2 after TGI provided significantly better survival rate than the administration immediately after TGI. Eventually, dosing twice on Day 2 and 5 exerted long-term therapeutic effects, which were confirmed by a Y-maze behavioral test demonstrating improved spatial memory compared with untreated rats on Day 20. Immunohistochemical analysis showed that astrocytes were chief targets of the BDNF mRNA-loaded nanomicelles, suggesting that the augmented BDNF secretion from astrocytes creates a supportive microenvironment for the neurons to tolerate changes caused by ischemic stresses, and terminate the process of progressive neuronal death after the ischemic attack. Overall, the unique mechanism of action of mRNA therapeutics provide a promising approach for preventing ischemic neuronal death.

PGE2 and Thrombin Induce Myofibroblast Transdifferentiation via Activin A and CTGF in Endometrial Stromal Cells.

Endometriosis is characterized by inflammation and fibrotic changes. Our previous study using a mouse model showed that proinflammatory factors present in peritoneal hemorrhage exacerbated inflammation in endometriosis-like grafts, at least in part through the activation of prostaglandin (PG) E2 receptor and protease-activated receptor (PAR). In addition, menstruation-related factors, PGE2 and thrombin (P/T), a PAR1 agonist induced epithelial-mesenchymal transition (EMT) of endometrial cells under hypoxia. However, the molecular mechanisms by which P/T induce development of endometriosis have not been fully characterized. To investigate the effects of P/T, RNA extracted from endometrial stromal cells (ESCs) treated with P/T were subjected to RNA sequence analysis, and identified activin A, FOS, and GATA2 as upregulated genes. Activin A increased the expression of connective tissue growth factor (CTGF) and mesenchymal marker genes in ESCs. CTGF induced the expression of fibrosis marker type I collagen, fibronectin, and α-smooth muscle actin (αSMA), indicating fibroblast to myofibroblast transdifferentiation (FMT) of ESCs. In addition, activin A, FOS, GATA2, CTGF, and αSMA were localized in endometriosis lesions. Taken together, our data show that P/T induces changes resembling EMT and FMT in ectopic ESCs derived from retrograde menstruation, and that these are associated with fibrotic changes in the lesions. Pharmacological means that block P/T-induced activin A and CTGF signaling may be strategies to inhibit fibrosis in endometriotic lesions.

Enhancement of Motor Function Recovery after Spinal Cord Injury in Mice by Delivery of Brain-Derived Neurotrophic Factor mRNA.

Spinal cord injury (SCI) is a debilitating condition that can cause impaired motor function or full paralysis. In the days to weeks following the initial mechanical injury to the spinal cord, inflammation and apoptosis can cause additional damage to the injured tissues. This secondary injury impairs recovery. Brain-derived neurotrophic factor is a secreted protein that has been shown to improve a variety of neurological conditions, including SCI, by promoting neuron survival and synaptic plasticity. This study treated a mouse model of contusion SCI using a single dose of brain-derived neurotrophic factor (BDNF) mRNA nanomicelles prepared with polyethylene glycol polyamino acid block copolymer directly injected into the injured tissue. BDNF levels in the injured spinal cord tissue were approximately doubled by mRNA treatment. Motor function was monitored using the Basso Mouse Scale and Noldus CatWalk Automated Gait Analysis System for 6 weeks post-injury. BDNF-treated mice showed improved motor function recovery, demonstrating the feasibility of mRNA delivery to treat SCI.

Cerebral Hyperperfusion Syndrome After a Burr Hole Drainage Surgery for Chronic Subdural Hematoma.

BACKGROUND: Although chronic subdural hematoma (CSDH) has a good prognosis after classical minimally-invasive drainage surgery, severe complications still occur at a substantial rate. Cerebral hyperperfusion syndrome (CHS), which is a common severe complication after carotid endarterectomy or carotid artery stenting for cervical carotid artery stenosis, is rare after drainage surgery for a CSDH. CASE DESCRIPTION: We describe the case of an 82-year-old woman who presented with ipsilesional symptoms including contralateral hemiparesis and dysarthria, progressively worsening consciousness, and status epilepticus after a burr hole drainage surgery for CSDH. Magnetic resonance fluid-attenuated inversion recovery imaging showed diffuse subcortical low intensity in the ipsilesional hemisphere almost simultaneously with the appearance of the symptoms. Arterial spin labeling magnetic resonance perfusion imaging showed the abnormal increase of cerebral blood flow in the hemisphere. Continuous propofol administration and blood pressure management improved the symptoms. CONCLUSIONS: CHS can cause severe postoperative complications after drainage surgery for CSDH. Subcortical low-intensity fluid-attenuated inversion recovery imaging is a useful investigation for early detection of CHS in CSDH, and arterial spin labeling imaging is an effective minimally-invasive modality for confirming the diagnosis.

Identification of Spring Coefficient for Heel Rocker Function Support Based on Estimated Dorsiflexion Torque.

In previous research, we have developed a high-dorsiflexion assistive robotic technology aiming for gait rehabilitation targeting on ankle dorsiflexion movement. A McKibben-type artificial muscle is applied to provide large dorsiflexion force while adding little weight to the device. This ensures the foot clearance before initial stance phase in gait. Meanwhile, a tension spring is deployed in series with the artificial muscle to support heel rocker function in loading response phase. Suitable spring coefficient for each individual differs according to ankle's dorsiflexion torque in loading response. An unsuitable spring would lead to knee deviation in this phase. In this study, we derived an identification equation to determine a suitable spring coefficient for individuals based on estimation of dorsiflexion torque required to support. An evaluation test on healthy objects was conducted, which shows no negative effects on participants' knee angles with the identified spring coefficient.

Histological evaluation of tissue damage caused by rotational needle insertion.

Needles used in percutaneous insertion must be as thin as possible to minimize invasiveness. However, using extra-thin needles with a diameter less than 25G (0.53 mm diameter) can cause needle deflection. Needle deflection can be minimized by insertion with axial rotation along the needle shaft; this rotation is also useful for steering the insertion direction of the needle tip. However, although high rotation speeds may decrease needle deflection, this may increase tissue damage. Therefore, the purpose of this study was to histologically evaluate tissue damage caused by the rotational needle-insertion method, and to verify the needle-tip deflection caused by tissue damage. In this paper, we evaluated tissue damage and needle deflection caused by needle insertion with no rotation, unidirectional rotation, and bidirectional rotation. The results suggest that percutaneous needle insertion under unidirectional rotation is potentially risky in humans, as this causes wound-up tissue and expansion of the area of the hole created by the needle path. In contrast, needle insertion under bidirectional rotation appeared to minimize deflection, and prevented winding of tissue and expansion of the hole created by the needle path.

Extremely Delayed Multiple Brain Metastases from Renal Cell Carcinoma: Remission Achieved with Total Surgical Removal: Case Report and Literature Review.

BACKGROUND: Late brain metastasis from renal cell carcinoma (RCC), which is generally considered as metastasis occurring more than 10 years after nephrectomy, often occurs as a solitary lesion, and total resection is recommended to achieve remission. CASE DESCRIPTION: We describe a rare case of multiple late brain metastases from RCC in a 60-year-old man who presented with 3 brain metastases from RCC 22 years after nephrectomy. Total removal of the 3 lesions achieved remission without adjuvant therapy. CONCLUSIONS: Total removal of late brain metastasis from RCC, even occurring with multiple lesions, can achieve total remission under specific conditions.

Repeated de novo aneurysm formation after anastomotic surgery: Potential risk of genetic variant RNF213 c.14576G>A.

BACKGROUND: De novo aneurysm formation after intracranial anastomotic surgery is a relatively rare complication with fewer than 20 reported cases, and the mechanism is still unclear. CASE DESCRIPTION: A 63-year-old male treated for symptomatic internal carotid artery occlusion developed de novo aneurysms twice after anastomoses first of the superficial temporal artery-middle cerebral artery and second of the external carotid artery-radial artery-middle cerebral artery over a 10-year period. The first de novo aneurysm was successfully resected with pathological diagnosis of true aneurysm. The second de novo aneurysm thrombosed naturally after gradual growth. Genetic testing of the patient revealed the c.14576G>A (p.R4859K) variant in ring finger protein 213, which is a susceptibility gene for moyamoya disease. CONCLUSIONS: This genetic variant was probably involved in the repeated de novo aneurysm formation, and this case represents a rare phenotype of the genetic variant.

Ptosis as partial oculomotor nerve palsy due to compression by infundibular dilatation of posterior communicating artery, visualized with three-dimensional computer graphics: case report.

Oculomotor nerve palsy (ONP) due to internal carotid-posterior communicating artery (PcomA) aneurysm generally manifests as partial nerve palsy including pupillary dysfunction. In contrast, infundibular dilatation (ID) of the PcomA has no pathogenic significance, and mechanical compression of the cranial nerve is extremely rare. We describe a 60-year-old woman who presented with progressive ptosis due to mechanical compression of the oculomotor nerve by an ID of the PcomA. Three-dimensional computer graphics (3DCG) accurately visualized the mechanical compression by the ID, and her ptosis was improved after clipping of the ID. ID of the PcomA may cause ONP by mechanical compression and is treatable surgically. 3DCG are effective for the diagnosis and preoperative simulation.

Effect of dural detachment on long-term tumor control for meningiomas treated using Simpson grade IV resection.

OBJECT: Meningiomas treated by subtotal or partial resection are associated with significantly shorter recurrence-free survival than those treated by gross-total resection. The Simpson grading system classifies incomplete resections into a single category, namely Simpson Grade IV, with wide variations in the volume and location of residual tumors, making it complicated to evaluate the achievement of surgical goals and predict the prognosis of these tumors. Authors of the present study investigated the factors related to necessity of retreatment and tried to identify any surgical nuances achievable with the aid of modern neurosurgical techniques for meningiomas treated using Simpson Grade IV resection. METHODS: This retrospective analysis included patients with WHO Grade I meningiomas treated using Simpson Grade IV resection as the initial therapy at the University of Tokyo Hospital between January 1995 and April 2010. Retreatment was defined as reresection or stereotactic radiosurgery due to postoperative tumor growth. RESULTS: A total of 38 patients were included in this study. Regrowth of residual tumor was observed in 22 patients with a mean follow-up period of 6.1 years. Retreatment was performed for 20 of these 22 tumors with regrowth. Risk factors related to significantly shorter retreatment-free survival were age younger than 50 years (p = 0.006), postresection tumor volume of 4 cm(3) or more (p = 0.016), no dural detachment (p = 0.001), and skull base location (p = 0.016). Multivariate analysis revealed that no dural detachment (hazard ratio [HR] 6.42, 95% CI 1.41-45.0; p = 0.02) and skull base location (HR 11.6, 95% CI 2.18-218; p = 0.002) were independent risk factors for the necessity of early retreatment, whereas postresection tumor volume of 4 cm(3) or more was not a statistically significant risk factor. CONCLUSIONS: Compared with Simpson Grade I, II, and III resections, Simpson Grade IV resection includes highly heterogeneous tumors in terms of resection rate and location of the residual mass. Despite the difficulty in analyzing such diverse data, these results draw attention to the favorable effect of dural detachment (instead of maximizing the resection rate) on long-term tumor control. Surgical strategy with an emphasis on detaching the tumor from the affected dura might be another important option in resection of high-risk meningiomas not amenable to gross-total resection.

Peer attachment formation by systemic redox regulation with social training after a sensitive period.

Attachment formation is the most pivotal factor for humans and animals in the growth and development of social relationships. However, the developmental processes of attachment formation mediated by sensory-motor, emotional, and cognitive integration remain obscure. Here we developed an animal model to understand the types of social interactions that lead to peer-social attachment formation. We found that the social interaction in a sensitive period was essential to stabilise or overwrite the initially imprinted peer affiliation state and that synchronised behaviour with others based on common motivations could be a driver of peer social attachment formation. Furthermore, feeding experience with supplementation of ubiquinol conferred peer social attachment formation even after the sensitive period. Surprisingly, the experience of feeding beyond the cage window was also effective to reduce the required amount ubiquinol, suggesting that peri-personal space modulation may affect socio-emotional cognition and there by lead to attachment formation.

A cross-species socio-emotional behaviour development revealed by a multivariate analysis.

Recent progress in affective neuroscience and social neurobiology has been propelled by neuro-imaging technology and epigenetic approach in neurobiology of animal behaviour. However, quantitative measurements of socio-emotional development remains lacking, though sensory-motor development has been extensively studied in terms of digitised imaging analysis. Here, we developed a method for socio-emotional behaviour measurement that is based on the video recordings under well-defined social context using animal models with variously social sensory interaction during development. The behaviour features digitized from the video recordings were visualised in a multivariate statistic space using principal component analysis. The clustering of the behaviour parameters suggested the existence of species- and stage-specific as well as cross-species behaviour modules. These modules were used to characterise the behaviour of children with or without autism spectrum disorders (ASDs). We found that socio-emotional behaviour is highly dependent on social context and the cross-species behaviour modules may predict neurobiological basis of ASDs.