Potential application of let-7a antagomir in injured peripheral nerve regeneration

Neurotrophic factors,particularly nerve growth factor,enhance neuronal regeneration.However,the in vivo applications of nerve growth factor are largely limited by its intrinsic disadvantages,such as its short biological half-life,its contribution to pain response,and its inability to cross the blood-brain barrier.Considering that let-7(human miRNA)targets and regulates nerve growth factor,and that let-7 is a core regulator in peripheral nerve regeneration,we evaluated the possibilities of let-7 application in nerve repair.In this study,anti-let-7a was identified as the most suitable let-7 family molecule by analyses of endogenous expression and regulatory relationship,and functional screening.Let-7a antagomir demonstrated biosafety based on the results of in vivo safety assessments and it entered into the main cell types of the sciatic nerve,including Schwann cells,fibroblasts and macrophages.Use of hydrogel effectively achieved controlled,localized,and sustained delivery of let-7a antagomir.Finally,let-7a antagomir was integrated into chitosan conduit to construct a chitosan-hydrogel scaffold tissue-engineered nerve graft,which promoted nerve regeneration and functional recovery in a rat model of sciatic nerve transection.Our study provides an experimental basis for potential in vivo application of let-7a.

S6B-2 Matrine Inhibits Itching by Lowering the Activity of Calcium Channel

Sophorae Flavescentis Radix(SFR)is a medicinal herb with many functions that are involved in anti-inflammation,antinociception,and anticancer.SFR is also used to treat a variety of itching diseases.Matrine(MT)is one of the main constituents in SFR and also has the effect of relieving itching,but the antipruritic mechanism is still unclear.Here,we investigated the effect of MT on antipruritus.In acute and chronic itch models,MT significantly inhibited the scratching behavior not only in acute itching induced by histamine(His),chloroquine(CQ)and compound 48/80 with a dose-depended manner,but also in the chronic pruritus models of Atopic dermatitis(AD)and Acetone-ether-water(AEW)in mice.Furthermore,MT can be detected in the blood after intraperitoneal injection(i.p.)and subcutaneous injection(s.c.).Finally,electrophysiological and calcium image results show that MT inhibits the excitatory synaptic transmission from dorsal root ganglion(DRG)to the dorsal horn of the spinal cord by suppressing presynaptic N-type calcium channels.Taken together,we believe that MT is a novel drug candidate in treating pruritus diseases,especially for histamine-independent and chronic pruritus,which might be attributed to inhibition of presynaptic N-type calcium channels.

Deficiency of transmembrane AMPA receptor regulatory protein γ-8 leads to attention-deficit hyperactivity disorder-like behavior in mice

Attention-deficit hyperactivity disorder(ADHD) is a neurodevelopmental disorder prevalent in schoolage children. At present, however, its etiologies and risk factors are unknown. Transmembrane α-amino-3-hydroxy-5-methyl-4-isoxazolepropionicacid(AMPA) receptor regulatory protein γ-8(TARP γ-8,also known as calcium voltage-gated channel auxiliary subunit gamma 8(CACNG8)) is an auxiliary AMPA receptor(AMPAR) subunit. Here, we report an association between TARP γ-8 and ADHD,whereby adolescent TARP γ-8 knockout(KO) mice exhibitedADHD-likebehaviors,including hyperactivity, impulsivity, anxiety, impaired cognition,and memory deficits. Human single-nucleotide polymorphism(SNP) analysis also revealed strong associations between intronic alleles in CACNG8genes and ADHD susceptibility. In addition,synaptosomal proteomic analysis revealed dysfunction of the AMPA glutamate receptor complex in the hippocampi of TARP γ-8 KO mice.Proteomic analysis also revealed dysregulation of dopaminergic and glutamatergic transmissions in the prefrontal cortices of TARP γ-8 KO mice.Methylphenidate(MPH), which is commonly used to treat ADHD, significantly rescued the major behavioral deficits and abnormal synaptosomal proteins in TARP γ-8 KO mice. Notably, MPH significantly reversed the up-regulation of Grik2 and Slc6a3 in the prefrontal cortex. MPH also significantly improved synaptic AMPAR complex function by up-regulating other AMPAR auxiliary proteins in hippocampal synaptosomes. Taken together, our results suggest that TARP γ-8 is involved in the development of ADHD in humans.This study provides a useful alternative animal model with ADHD-like phenotypes related to TARP γ-8deficiency, which has great potential for the development of new therapies.

肝胆肿瘤中肿瘤特异性CircRNA衍生抗原肽的鉴定

基于肿瘤抗原的免疫治疗的应用受到验证免疫原性肽稀缺性的阻碍。本研究旨在研究环状RNA(circRNA)在肝胆肿瘤类器官中作为肿瘤抗原肽新来源的潜力。使用RNA测序(RNA-seq)和基于算法的评分工具,预测3950个翻译的肿瘤特异性环状RNA在27个类器官中产生18971个抗原肽。从抗原格局来看,11个氨基酸长度(mer)肽和人白细胞抗原(HLA)-A结合肽具有最高的免疫原性相关评分。在分析的3/5类器官中,有13个预测抗原肽通过质谱(MS)免疫肽组学被直接确认为HLA-A、HLA-B和HLA-C(HLA-ABC)结合肽。在流式细胞术和酶联免疫吸附试验(ELISA)中,由HLA-ABC分子呈递的circRNA衍生的肿瘤特异性肽刺激CD8(CD8)T细胞,显示CD107a干扰素γ(IFNγ)共表达和IFNγ分泌增加。免疫原性环状RNA衍生肽诱导的靶向类器官的细胞毒性T细胞活性在杀伤实验中得到验证。值得注意的是,来自circTBC1D15的抗原肽YGFNEILKK不仅被认为是类器官的HLA-ABC呈递肽,而且还显著降低了肿瘤类器官的存活率。本研究的发现强调了产生肿瘤抗原的一个关键亚群,这对靶向肿瘤特异性circRNA具有重要意义。

A bacterial artificial chromosome transgenic mouse model for visualization of neurite growth

Class III β-tubulin(Tubb3) is a component of the microtubules in neurons and contributes to microtubule dynamics that are required for axon outgrowth and guidance during neuronal development. We here report a novel bacterial artificial chromosome(BAC) transgenic mouse line that expresses Class III β-tubulin fused to m Cherry, an improved monomeric red fluorescent protein, for the visualization of microtubules during neuronal development. A BAC containing Tubb3 gene was modified by insertion of m Cherry complementary DNA downstream of Tubb3 coding sequence via homologous recombination. m Cherry fusion protein was expressed in the nervous system and testis of the transgenic animal, and the fluorescent signal was observed in the neurons that located in the olfactory bulb, cerebral cortex, hippocampal formation, cerebellum, as well as the retina. Besides, Tubb3-m Cherry fusion protein mainly distributed in neurites and colocalized with endogenous Class III β-tubulin. The fusion protein labels Purkinje cell dendrites during cerebellar circuit formation. Therefore, this transgenic line might be a novel tool for scientific community to study neuronal development both in vitro and in vivo.

A β-sheet-targeted theranostic agent for diagnosing and preventing aggregation of pathogenic peptides in Alzheimer’s disease

Amyloid-βpeptide(Aβ)aggregates,particularly Aβoligomers,are established biomarker and toxic species in Alzheimer’s disease(AD).Early detection and disaggregation of Aβaggregates are of great importance for the treatment of AD due to the unavailability of therapy at the advanced stages of the disease.A multitalented agent,2-{2-[(1 H-benzoimidazol-2-yl)methoxy]phenyl}benzothiazole(BPB),is designed by merging twoβ-sheet targeting groups into one molecule to detect and inhibit the Aβaggregation.BPB can quantitatively measure theβ-sheet level of soluble Aβoligomers and specifically distinguish the aggregates of Aβ40 and Aβ42 by unique luminescence spectrum.Animal tests demonstrate that BPB can efficiently penetrate the blood brain barrier and precisely stain Aβplaques in the brain;more importantly,it can differentiate the blood of APP transgenic mice from that of normal ones.In addition to the diagnostic potential,BPB also suppresses the generation of ROS,protects the neurons from neurotoxicity,and disaggregates the Aβaggregates in brain homogenates of APP transgenic mice induced by metal ions or self-assembly.In view of its detective ability toward Aβoligomers and inhibition to Aβ-related neurotoxicity,BPB may be developed into a sensitive probe for screening blood samples in the early diagnosis of AD as well as an effective inhibitor for diminishing Aβaggregates in the treatment of the disease.

TMEM63B regulates postnatal development of cochlear sensory epithelia via thyroid hormone signaling

Sound transmission occurs in the cochlea,a complex and ingenious subdivision in the inner ear.The structure of the cochlea develops structurally and functionally by the time before two postnatal weeks(the time of hearing onset)in mice(Geal-Dor et al.,1993).Greater epithelial ridge(GER,also known as Kolliker's organ)is a transient cochlear structure containing a group of columnar epithelial supporting cells surrounding the inner hair cells(IHCs).

The characterization of protein lactylation in relation to cardiac metabolic reprogramming in neonatal mouse hearts

In mammals,the neonatal heart can regenerate upon injury within a short time after birth,while adults lose this ability.Metabolic reprogramming has been demonstrated to be critical for cardiomyocyte proliferation in the neonatal heart.Here,we reveal that cardiac metabolic reprogramming could be regulated by altering global protein lactylation.By performing 4D label-free proteomics and lysine lactylation(Kla)omics analyses in mouse hearts at postnatal days 1,5,and 7,2297 Kla sites from 980 proteins are identified,among which 1262 Kla sites from 409 proteins are quantified.Functional clustering analysis reveals that the proteins with altered Kla sites are mainly involved in metabolic processes.The expression and Kla levels of proteins in glycolysis show a positive correlation while a negative correlation in fatty acid oxidation.Furthermore,we verify the Kla levels of several differentially modified proteins,including ACAT1,ACADL,ACADVL,PFKM,PKM,and NPM1.Overall,our study reports a comprehensive Kla map in the neonatal mouse heart,which will help to understand the regulatory network of metabolic reprogramming and cardiac regeneration.

Model animals and their applications

Looking back a decade ago when I decided to use"model animal"to name our newly established institute,my outrageous boldness can only be described by the old Chinese slang,"newborn calf could never sense the danger of tiger".

Protein prenylation and human diseases: a balance of protein farnesylation and geranylgeranylation

The protein prenylation is one of the essential post-translational protein modifications, which extensively exists in the eukaryocyte. It includes protein farnesylation and geranylgeranylation, using farnesyl pyrophosphate(FPP) or geranylgeranyl pyrophosphate(GGPP) as the substrate, respectively. The prenylation occurs by covalent addition of these two types of isoprenoids to cysteine residues at or near the carboxyl terminus of the proteins that possess Caa X motif, such as Ras small GTPase family. The attachment of hydrophobic prenyl groups can anchor the proteins to intracellular membranes and trigger downstream cell signaling pathway. Geranylgeranyl biphosphate synthase(GGPPS) catalyzes the synthesis of 20-carbon GGPP from 15-carbon FPP. The abnormal expression of this enzyme will affect the relative content of FPP and GGPP, and thus disrupts the balance between protein farnesylation and geranylgeranylation, which participates into various aspects of cellular physiology and pathology. In this paper, we mainly review the property of this important protein post-translational modification and research progress in its regulation of cigarette smoke induced pulmonary disease, adipocyte insulin sensitivity, the inflammation response of Sertoli cells, the hepatic lipogenesis and the cardiac hypertrophy.

Towards a better understanding of mouse and human diseases—International Mouse Phenotyping Consortium

With the completion of the human genome sequence,the last several decades have seen the pursuit of gene function in vivo.The mouse is generally considered the best model animal for examining gene function because of its high productivity,pure genetic background,and degree of similarity to the human genome.With the development of sev-

Corrigendum to“Actin polymerization induces mitochondrial distribution during collective cell migration”[Journal of Genetics and Genomics(2023)50 46-49]

This erratum clarifies information in the Letter to the Editor“Actin polymerization induces mitochondrial distribution during collective cell migration”by Qu et al.(2023).In the section for the list of author names,“Chen Qu,Yating Kan,Hui Zuo,Mengqi Wu,Zhixiang Dong,Xinyi Wang,Qing Zhang,Heng Wang,Dou Wang,Jiong Chen”should be“Chen Qu,Yating Kan,Xinyi Wang,Hui Zuo,Mengqi Wu,Zhixiang Dong,Qing Zhang,Heng Wang,Dou Wang,Jiong Chen”.

Deficiency of Autophagy-Related Gene 5 in Keratinocytes Leads to Aggravation of Epidermal Damage in 2,4-Dinitrochlorobenzene-Induced Allergic Contact Dermatitis

Objective:The interrelationship between apoptosis and autophagy plays an important role in many pathophysiological processes,however,whether their interplay is involved in allergic contact dermatitis(ACD)has not yet been elucidated.So,we conducted this study to determine whether keratinocyte-specific autophagy-related gene 5(ATG5)deficiency can regulate apoptosis to inhibit skin damage in mice with 2,4-dinitrochlorobenzene(DNCB)-induced ACD.Methods:This study involved keratinocyte-specificAtg5 conditional knockout(cKO)mice(Krt14cre/+-Atg5flox/flox)and control mice(Krt14+/+-Atg5flox/flox).We painted DNCB on the right ear of each mouse to induce ACD.Dermatitis scoring and measurements of ear weight and thickness were performed to evaluate inflammation levels.An immunohistochemical assay was performed to analyze immune cell infiltration.Histological study and TUNEL staining were performed to compare the differences in skin lesions betweenAtg5 cKO mice and control mice.Immunofluorescence and western blotting were used to examine the levels of ATG5 and apoptosis-related protein.The results were statistically analyzed byt test.Results:After DNCB stimulation of mice ears,we observed a more severe phenotype inAtg5 cKO mice than in control mice(dermatitis score:7.500±2.588vs.3.250±0.822,P=0.003).Further analysis of ATG5 protein confirmed keratinocyte-specific ablation ofAtg5 in cKO mice and showed that DNCB did not influence ATG5 expression.Immunohistochemistry assay revealed that the infiltrated immune cells were not involved in aggravation of the phenotype of DNCB-stimulatedAtg5 cKO mice.However,the histological study(P=0.024),TUNEL staining(P=0.024),immunofluorescence(P=0.036),and western blotting showed that the increase in keratinocyte death,especially apoptosis,contributed to aggravation of the phenotype of DNCB-stimulatedAtg5 cKO mice.Conclusion:Deficiency ofAtg5 in keratinocytes increases apoptosis,aggravating skin damage in DNCB-induced ACD mice.This has no relationship with the involvement of immune cells.

LonP1 Links Mitochondria–ER Interaction to Regulate Heart Function

Interorganelle contacts and communications are increasingly recognized to play a vital role in cellular function and homeostasis.In particular,the mitochondria–endoplasmic reticulum(ER)membrane contact site(MAM)is known to regulate ion and lipid transfer,as well as signaling and organelle dynamics.However,the regulatory mechanisms of MAM formation and their function are still elusive.Here,we identify mitochondrial Lon protease(LonP1),a highly conserved mitochondrial matrix protease,as a new MAM tethering protein.The removal of LonP1 substantially reduces MAM formation and causes mitochondrial fragmentation.Furthermore,deletion of LonP1 in the cardiomyocytes of mouse heart impairs MAM integrity and mitochondrial fusion and activates the unfolded protein response within the ER(UPR^(ER)).Consequently,cardiac-specific LonP1 deficiency causes aberrant metabolic reprogramming and pathological heart remodeling.These findings demonstrate that LonP1 is a novel MAM-localized protein orchestrating MAM integrity,mitochondrial dynamics,and UPR^(ER),offering exciting new insights into the potential therapeutic strategy for heart failure.

Molecular mechanisms of dietary restriction in aging—insights from Caenorhabditis elegans research

Dietary restriction(DR) is one of the most robust environmental manipulations that not only extend life span but also delay the onset of age-related diseases in almost every species examined. Caenorhabditis elegans plays an important role in aging studies due to its simple life cycle, easy genetic manipulations and highly conserved genome. Recent studies have demonstrated that the beneficial effects of DR are mediated by the highly conserved transcription factors and signaling pathways in C. elegans. Here we review recent progress in the methodology and molecular mechanisms of DR using C. elegans as a model, as well as prospects for future research.

Inactivation of Cipc alters the expression of Per1 but not circadian rhythms in mice

Circadian clocks are comprised of self-sustained transcriptional/translational feedback loops, which regulate the rhythms of physiology and behavior in mammals. CLOCK-interacting protein, Circadian(CIPC), has been indicated as an additional negative-feedback regulator of the circadian clock in vitro, although its physiological roles in circadian clock are unknown. Here, we generated Cipc homozygous knockout(Cipc-/-) mice and assessed the resultant circadian phenotypes. Surprisingly, the m RNA expression profiles of core clock genes in the liver of Cipc-/- mice showed no significant differences from that in wild-type mice except for Per1. Cipc-/- mice displayed normal locomotor rhythm and entrained activity pattern in both 12:12 light-dark cycle and constant dark cycle. Furthermore, deletion of Cipc in lungs and adipose tissues did not influence their peripheral clock. The results from this work provided more conclusive data suggesting that CIPC is not critically required for basic clock function.

Germ layer formation during Xenopus embryogenesis: the balance between pluripotency and differentiation

The African clawed frog, Xenopus laevis, has long been a model animal for the studies in the fields of animal cloning, developmental biology, biochemistry, cell biology, and physiology. With the aid of Xenopus, major molecular mechanisms that are involved in embryonic development have been understood. Germ layer formation is the first event of embryonic cellular differentiation, which is induced by a few key maternal factors and subsequently by zygotic signals. Meanwhile, another type of signals, the pluripotency factors in ES cells, which maintain the undifferentiated state, are also present during early embryonic cells. In this review, the functions of the pluripotency factors during Xenopus germ layer formation and the regulatory relationship between the signals that promote differentiation and pluripotency factors are discussed.

Generation of gene-target dogs using CRISPR/Cas9 system

Dear Editor,Dogs(Canis familiaris)serve as human companions and are raised to herd livestock,aid hunters,guard homes,perform police and rescue work,and guide the blind.Dogs exhibit close similarities to humans in terms of metabolic,physiological,and anatomical characteristics,and thus are ideal genetic and clinical models to study human diseases(Tsai et al.,2007).Gene target technology is a powerful tool to create new strains of animals with favorable traits.However,thus far,gene-target dogs have not been developed due to their unique species-specific reproductive characteristics,which limits the applications of dogs especially in the field of biomedical research.Recently,clustered regularly interspaced short palindromic repeats(CRISPRs)/CRISPR-associated(Cas)9 system was applied to edit specific genes with a high efficiency(Cong et al.,2013;Mali et al.,2013).Here we attempt to explore the feasibility of producing gene knockout(KO)dogs by using this technology.Beagle dog,the most widely used breed in biomedical research,was used as our animal model.Myostatin(MSTN)was chosen as the first gene of interest.

Glucose metabolic abnormality is associated with defective mineral homeostasis in skeletal disorder mouse model

Bone was reported as a crucial organ for regulating glucose homeostasis. In this study, we found that Phex mutant mice(PUG), a model of human X-linked hypophosphatemic rickets(XLH), displayed metabolic abnormality in addition to abnormal phosphate homeostasis, skeletal deformity and growth retardation. Glucose tolerance was elevated with enhanced insulin sensitivity in PUG, though circulating insulin level decreased. Interestingly, bone mineral density defects and glucose metabolic abnormality were both rescued by adding phosphorus- and calcium-enriched supplements in daily diet. Serum insulin level, glucose tolerance and insulin sensitivity showed no differences between PUG and wild-type mice with rescued osteocalcin(OCN) following treatment. Our study suggested that OCN is a potential mediator between mineral homeostasis and glucose metabolism. This investigation brings a new perspective on glucose metabolism regulation through skeleton triggered mineral homeostasis and provides new clues in clinical therapeutics of potential metabolic disorders in XLH patients.

In vivo RNAi screen identifies candidate signaling genes required for collective cell migration in Drosophila ovary

Collective migration of loosely or closely associated cell groups is prevalent in animal development, physiological events, and cancer metastasis. However, our understanding of the mechanisms of collective cell migration is incomplete. Drosophila border cells provide a powerful in vivo genetic model to study collective migration and identify essential genes for this process. Using border cell-specific RNAi-silencing in Drosophila, we knocked down 360 conserved signaling transduction genes in adult flies to identify essential pathways and genes for border cell migration. We uncovered a plethora of signaling genes, a large proportion of which had not been reported for border cells, including Rack1(Receptor of activated C kinase) and brk(brinker), mad(mother against dpp), and sax(saxophone), which encode three components of TGF-β signaling. The RNAi knock down phenotype was validated by clonal analysis of Rack1 mutants. Our data suggest that inhibition of Src activity by Rack1 may be important for border cell migration and cluster cohesion maintenance. Lastly, results from our screen not only would shed light on signaling pathways involved in collective migration during embryogenesis and organogenesis in general, but also could help our understanding for the functions of conserved human genes involved in cancer metastasis.