Effects of fluorescent protein tdTomato on mouse retina.

Fluorescent proteins (FPs) have been widely used to investigate cellular and molecular interactions and trace biological events in many applications. Some of the FPs have been demonstrated to cause undesirable cellular damage by light-induced ROS production in vivo or in vitro. However, it remains unknown if one of the most popular FPs, tdTomato, has similar effects in neuronal cells. In this study, we discovered that tdTomato expression led to unexpected retinal dysfunction and ultrastructural defects in the transgenic mouse retina. The retinal dysfunction mainly manifested in the reduced photopic electroretinogram (ERG) responses and decreased contrast sensitivity in visual acuity, caused by mitochondrial damages characterized with cellular redistribution, morphological modifications and molecular profiling alterations. Taken together, our findings for the first time demonstrated the retinal dysfunction and ultrastructural defects in the retinas of tdTomato-transgenic mice, calling for a more careful design and interpretation of experiments involved in FPs.

Hsa-miR-877-5p Expression in Acute Ischemic Stroke Based on Bioinformatics Analysis and Clinical Validation.

Inflammation and immunity play important roles in the pathogenesis of ischemic stroke. This study aimed to explore key regulatory genes in acute ischemic stroke (AIS) and their underlying mechanisms to provide new research targets for the diagnosis and treatment of ischemic stroke. We searched for differentially expressed mRNAs and miRNAs in patients with AIS and healthy populations in GEO databases, constructed a miRNA-mRNA network, and screened key miRNAs using least absolute shrinkage and selection operator regression and the support vector machine-recursive feature elimination model. Correlations between key miRNAs and infiltrating immune cells and inflammatory factors were analyzed using CIBERSORT and immunoassays and verified using clinical experiments. Bioinformatics analysis identified hsa-miR-877-5p as a key regulatory miRNA in AIS that can modulate immune and inflammatory responses. In clinical studies, it was verified by quantitative PCR analysis that the expression of hsa-miR-877-5p in the blood of AIS patients was higher than that of the healthy group. Then, enzyme-linked immunosorbent assay revealed that the expression of IL-23 and TNF-α related to inflammation in AIS patients was higher than that of the healthy. Quantitative PCR further found that the relative mRNA expression of IL-23, CXCR3, and TNF-α in AIS group was higher than that of the healthy group. This study may provide a basis for a more comprehensive understanding of the potential mechanism of the occurrence and development of AIS, and hsa-miR-877-5p and its downstream effectors IL-23, CXCR3, and TNF-α may be potential intervention targets in AIS.

Rhabdomyolysis-induced acute kidney injury after administration of a red yeast rice supplement: A case report.

BACKGROUND: A few reports have revealed induction of rhabdomyolysis by a red yeast rice (RYR) supplement or by RYR in combination with abiraterone (an androgen biosynthesis inhibitor). CASE SUMMARY: A 76-year-old man presented with progressive limb weakness, muscle soreness, and acute kidney injury (AKI). He had been taking the anti-prostate cancer drug abiraterone for 14 mo and had added a RYR supplement 3 mo before symptom onset. After being diagnosed with rhabdomyolysis-induced AKI, the patient discontinued these drugs and responded well to hemodialysis and hemoperfusion. After 23 d of treatment, creatine kinase levels returned to normal and serum creatinine levels decreased. CONCLUSION: We speculate that statins, the main lipid-lowering component of RYR, or a combination of statins and abiraterone, will increase the risk of rhabdomyolysis.

Curcumin alleviates orofacial allodynia and improves cognitive impairment via regulating hippocampal synaptic plasticity in a mouse model of trigeminal neuralgia.

OBJECTIVE: Cognitive impairment, one of the most prevalent complications of trigeminal neuralgia, is troubling for patients and clinicians due to limited therapeutic options. Curcumin shows antinociception and neuroprotection pharmacologically, suggesting that it may have therapeutic effect on this complication. This study aimed to investigate whether curcumin alleviates orofacial allodynia and improves cognitive impairment by regulating hippocampal CA1 region synaptic plasticity in trigeminal neuralgia. METHODS: A mouse model of trigeminal neuralgia was established by partially transecting the infraorbital nerve (pT-ION). Curcumin was administered by gavage twice daily for 14 days. Nociceptive thresholds were measured using the von Frey and acetone test, and the cognitive functions were evaluated using the Morris water maze test. Dendritic spines and synaptic ultrastructures in the hippocampal CA1 area were observed by Golgi staining and transmission electron microscopy. RESULTS: Curcumin intervention increased the mechanical and cold pain thresholds of models. It decreased the escape latency and distance to the platform and increased the number of platform crossings and dwell time in the target quadrant of models, and improved spatial learning and memory deficits. Furthermore, it partially restored the disorder of the density and proportion of dendritic spines and the abnormal density and structure of synapses in the hippocampal CA1 region of models. CONCLUSION: Curcumin alleviates abnormal orofacial pain and cognitive impairment in pT-ION mice by a mechanism that may be related to the synaptic plasticity of hippocampal CA1, suggesting that curcumin is a potential strategy for repairing cognitive dysfunction under long-term neuropathic pain conditions.

Electroacupuncture alleviates orofacial allodynia and anxiety-like behaviors by regulating synaptic plasticity of the CA1 hippocampal region in a mouse model of trigeminal neuralgia.

Objective: Trigeminal neuralgia (TN), one of the most severe and debilitating chronic pain conditions, is often accompanied by mood disorders, such as anxiety and depression. Electroacupuncture (EA) is a characteristic therapy of Traditional Chinese Medicine with analgesic and anxiolytic effects. This study aimed to investigate whether EA ameliorates abnormal TN orofacial pain and anxiety-like behavior by altering synaptic plasticity in the hippocampus CA1. Materials and methods: A mouse infraorbital nerve transection model (pT-ION) of neuropathic pain was established, and EA or sham EA was used to treat ipsilateral acupuncture points (GV20-Baihui and ST7-Xiaguan). Golgi-Cox staining and transmission electron microscopy (TEM) were administrated to observe the changes of synaptic plasticity in the hippocampus CA1. Results: Stable and persistent orofacial allodynia and anxiety-like behaviors induced by pT-ION were related to changes in hippocampal synaptic plasticity. Golgi stainings showed a decrease in the density of dendritic spines, especially mushroom-type dendritic spines, in hippocampal CA1 neurons of pT-ION mice. TEM results showed that the density of synapses, membrane thickness of the postsynaptic density, and length of the synaptic active zone were decreased, whereas the width of the synaptic cleft was increased in pT-ION mice. EA attenuated pT-ION-induced orofacial allodynia and anxiety-like behaviors and effectively reversed the abnormal changes in dendritic spines and synapse of the hippocampal CA1 region. Conclusion: EA modulates synaptic plasticity of hippocampal CA1 neurons, thereby reducing abnormal orofacial pain and anxiety-like behavior. This provides evidence for a TN treatment strategy.

CLEC3B is a novel causative gene for macular-retinal dystrophy.

PURPOSE: Macular degeneration is the leading cause of blindness worldwide. In this study, we aimed to define a new subtype of macular-retinal dystrophy and its genetic predisposition in 5 families. METHODS: Exome sequencing was performed to determine the putative disease-causing genes in patients with inherited macular disorders confirmed through comprehensive ophthalmic examinations. To validate its functional consequence, adeno-associated virus-mediated mutant gene was delivered into the murine retina, and both structural and functional tests were performed to investigate its pathological effects in vivo. RESULTS: In total, 5 multigenerational families diagnosed with autosomal dominant maculoretinopathy were found to carry a pathogenic variant in a new gene, CLEC3B, which encodes tetranectin, a plasminogen kringle-4 binding protein. Consistent with the disease phenotypes of patients, mice that received subretinal injections with the CLEC3B variant displayed multiple subretinal hyperreflective deposits, reduced retinal thickness, and decreased electroretinographic responses. Moreover, the optokinetic tracking response indicated that spatial frequency was significantly lower (P < .05), implying impaired visual function in these mice. CONCLUSION: We have presented a new subtype of macular-retinal dystrophy in 5 families as well as a new pathogenic gene, CLEC3B, providing new insights into maculoretinopathy etiology.

A Resting-State Functional Magnetic Resonance Imaging Study of Abnormal Frontoparietal Network Function in Migraine without Aura.

BACKGROUND This study aimed to investigate frontoparietal network (FPN) dysfunction in participants with migraine without aura (MwoA). MATERIAL AND METHODS We selected 48 age-, sex-, and education level-matched graduate students (24 participants with MwoA [MwoA group] and 24 healthy controls). RS-fMRI and independent component analysis were used to examine the FPN and to compare abnormal encephalic regional homogeneity values. The Mindful Attention Awareness Scale (MAAS), Self-Rating Anxiety Scale (SAS), Self-Rating Depression Scale (SDS), and Self-Rating Scale of Sleep (SRSS) were used to evaluate attention, anxiety, depression, and sleep, respectively. Pearson's correlation was applied to evaluate the association between abnormal brain areas and the scores for each scale. RESULTS Neural function activity in encephalic regions of FPN showed abnormal changes in the MwoA group. The MwoA group had significantly lower MAAS scores (P<0.001), higher SAS scores (P<0.001), and higher SDS (P=0.06) and SRSS scores (P=0.26). In the MwoA group, functional activity of the right parietal lobule in the left FPN was positively correlated with MAAS scores (P=0.01) and negatively correlated with SAS (P=0.02). The orbital part of left inferior frontal gyrus activity in the right FPN was positively correlated with SDS (P=0.04) and SRSS (P<0.001). Right superior marginal gyrus activity in the right FPN was positively correlated with SDS (P=0.02). CONCLUSIONS Abnormal FPN function was correlated with attention, anxiety, depression, and sleep status in the MwoA group. These results offer further insights into the evaluation and treatment of MwoA.

Circular Rims2 Deficiency Causes Retinal Degeneration.

Circular RNAs (circRNAs) refer to a newly recognized family of non-coding RNA with single-stranded RNAs. Despite emerging evidence indicating that circRNAs are abundantly expressed in various tissues, especially in the brain and retina, the role of circRNAs in retinal function and diseases is still largely unknown. Circular Rims2 (circRims2) is highly expressed and conserved in both the human and mouse brains. However, little is known about the expression and function of circRims2 in the retina. In the current study, the high-throughput RNA-seq analysis reveals a high expression of circRims2 in the retina. In addition, it is found that circRims2 is mainly located in plexiform layers that contain synapses between retinal neurons. Knocking down circRims2 with short hairpin RNA through subretinal adeno-associated viral (AAV) delivery in the mice leads to the decrease of the thickness of the outer and inner segment (OS/IS) layers and outer nuclear layer (ONL), and cessation of scotopic and photopic electroretinogram responses. Furthermore, the current study finds that circRims2 deficiency evokes retinal inflammation and activates the tumor necrosis factor (TNF) signaling pathway. Therefore, circRims2 may play an important role in the maintenance of retinal structure and function, and circRims2 deficiency may lead to pathogenic changes in the retina.

Retinal Degeneration Caused by Ago2 Disruption.

Purpose: Argonaute proteins are key players in small RNA-guided gene silencing processes. Ago2 is the member of the Argonaute subfamily with slicer endonuclease activity and is critical for microRNA homeostasis and indispensable for biological development. However, the impact of Ago2 dysregulation in the retina remains to be fully explored. In this study, we studied the role of Ago2 in mouse retina. Methods: We explored the function of Ago2 in the mouse retina through an adeno-associated virus-mediated Ago2 disruption mouse model. An ERG was carried out to determine the retinal function. Spectral domain optical coherence tomography, fundus photographs, and immunostaining were performed to investigate the retinal structure. A quantitative RT-PCR assay was used to determine the expression of noncoding RNAs. Results: Both silencing and overexpression of Ago2 in mouse retina resulted in significant retinal morphological alterations and severe impairment of retinal function, mainly with a thinned outer nuclear layer, shortened inner segment/outer segment, and diminished ERG responses. Furthermore, Ago2 disruption resulted in alterations of noncoding RNAs in retina. Conclusions: Our finding demonstrated that Ago2 interruption led to severe retinal degeneration, suggested that Ago2 homeostasis contributed to retinal structural and functional maintenance.

Abundant Neural circRNA Cdr1as Is Not Indispensable for Retina Maintenance.

Cdr1as is the abundant circular RNA (circRNA) in human and vertebrate retinas. However, the role of Cdr1as in the retina remains unknown. In this study, we aimed to generate a Cdr1as knockout (KO) mouse model and investigate the retinal consequences of Cdr1as loss of function. Through in situ hybridization (ISH), we demonstrated that Cdr1as is mainly expressed in the inner retina. Using CRISPR/Cas9 targeting Cdr1as, we successfully generated KO mice. We carried out ocular examinations in the KO mice until postnatal day 500. Compared with the age-matched wild-type (WT) siblings, the KO mice displayed increased b-wave amplitude of photopic electrophysiological response and reduced vision contrast sensitivity. Through small RNA profiling of the retinas, we determined that miR-7 was downregulated, while its target genes were upregulated. Taken together, our results demonstrated for the first time that Cdr1as ablation led to a mild retinal consequence in mice, indicating that Cdr1as abundance is not indispensable for retinal development and maintenance.

Astrocytic CCAAT/Enhancer-binding protein delta contributes to reactive oxygen species formation in neuroinflammation.

Excessive reactive oxygen species (ROS) can form an oxidative stress and an associated neuroinflammation. However, the contribution of astrocytes to ROS formation, the cause of the resistance of astrocytes to oxidative stress, and the consequences on neurons remain largely uninvestigated. The transcription factor CCAAT/enhancer-binding protein delta (CEBPD) is highly expressed in astrocytes and has been suggested to contribute to the progress of Alzheimer's disease (AD). In this study, we found that ROS formation and expression of p47phox and p67phox, subunits of NADPH oxidase, were increased in AppTg mice but attenuated in AppTg/Cebpd-/- mice. Cebpd can up-regulate p47phox and p67phox transcription via a direct binding on their promoters, which results in an increase in intracellular oxidative stress. In addition, Cebpd also up-regulated Cu/Zn superoxide dismutase (Sod1) in astrocytes. Inactivation of Sod1 increased the sensitization to oxidative stress, which provides a reason for the resistance of astrocytes in an oxidative stress environment. Taken together, the study first revealed and dissected the involvement of astrocytic Cebpd in the promotion of oxidative stress and the contribution of CEBPD to the resistance of astrocytes in an oxidative stress environment.

An intranasally delivered peptide drug ameliorates cognitive decline in Alzheimer transgenic mice.

Alzheimer's disease (AD) is the most common neurodegenerative disease. Imbalance between the production and clearance of amyloid ß (Aß) peptides is considered to be the primary mechanism of AD pathogenesis. This amyloid hypothesis is supported by the recent success of the human anti-amyloid antibody aducanumab, in clearing plaque and slowing clinical impairment in prodromal or mild patients in a phase Ib trial. Here, a peptide combining polyarginines (polyR) (for charge repulsion) and a segment derived from the core region of Aß amyloid (for sequence recognition) was designed. The efficacy of the designed peptide, R8-Aß(25-35), on amyloid reduction and the improvement of cognitive functions were evaluated using APP/PS1 double transgenic mice. Daily intranasal administration of PEI-conjugated R8-Aß(25-35) peptide significantly reduced Aß amyloid accumulation and ameliorated the memory deficits of the transgenic mice. Intranasal administration is a feasible route for peptide delivery. The modular design combining polyR and aggregate-forming segments produced a desirable therapeutic effect and could be easily adopted to design therapeutic peptides for other proteinaceous aggregate-associated diseases.

Serum Levels of 25-hydroxyvitamin D and Functional Outcome in Older Patients with Hip Fracture.

The aim was to assess prognostic value of serum 25-hydroxyvitamin D (25[OH] D) levels in older Chinese patients with hip fracture. From June, 2012 to February, 2014, older patients with hip fracture were included. Serum 25(OH) D levels were measured at admission. The functional evaluation at the time of discharge was performed by the Barthel Index. In the 66 patients with an unfavorable outcome, serum 25(OH) D levels were lower compared with those with a favorable outcome. In multivariate analyses, there was an increased risk of unfavorable outcome associated with serum 25(OH) D levels ≤ 20 ng/ml (OR 5.25, 95% CI: 3.12-8.16). Our data supported an association between serum 25[OH] D levels at admission and short-term prognosis in Chinese older patients with hip fracture.

Osteogenic potential of human calcitonin gene-related peptide alpha gene-modified bone marrow mesenchymal stem cells.

BACKGROUND: Most of the basic and clinical studies of osteonecrosis of the femoral head (ONFH) are restricted to bone tissues only, whereas various systems are involved in the onset and development of ONFH, including nervous system. Peptidergic nerve participates in the neuronal regulation of bone metabolism and anabolism, and plays key roles in the growth, repair and reconstruction of bone. Calcitonin gene-related peptide (CGRP), which is secreted by peptidergic nerve, is the main mediator of bone metabolism. It dramatically promotes the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs). Additionally, it enhances the osteoblast mass and the rate of osteoblast formation, and reduces the bone resorption by acting on osteoblasts and osteoclasts. Hence, we aimed to construct recombinant retrovirus vector pLNCX(2)-hCGRPα and to investigate the proliferation and osteogenic potential of hCGRPα-producing BMSCs (BMSCs/pLNCX(2)-hCGRPα) after virus infection. METHODS: The constructed recombinant retrovirus vector pLNCX(2)-hCGRPα was transfected into PT67 packaging cells by lipofectamine 2000. Virus was collected for BMSCs infection. The mRNA and protein expression of hCGRPα was examined by reverse transcription polymerase chain reaction (RT-PCR) and Western blotting, respectively. The cell proliferation was determined by methyl thiazoleterazolium (MTT) assay. The osteogenic potential of BMSCs was evaluated by alkaline phosphatase (ALP) activity. RESULTS: Both mRNA and protein expression of hCGRPα was detected in BMSCs/pLNCX(2)-hCGRPα cells. These cells exhibited significantly elevated proliferation and ALP value as compared with control BMSCs (P < 0.05). CONCLUSION: BMSCs/pLNCX(2)-hCGRPα cells could stably express hCGRPα and showed promoted proliferation ability and osteogenic potential as compared with control BMSCs.