mTOR is a highly conserved serine/threonine protein kinase that is critical for diverse cellular processes in both developmental and physiological settings. mTOR interacts with a set of molecules ...including Raptor and Rictor to form two distinct functional complexes, namely the mTORC1 and mTORC2. Here, we used novel genetic models to investigate functions of the mTOR pathway for cranial neural crest cells (NCCs), which are a temporary type of cells arising from the ectoderm layer and migrate to the pharyngeal arches participating craniofacial development. mTOR deletion elicited a proliferation deficit and excessive apoptosis of post-migratory NCCs, leading to growth arrest of the facial primordia along with midline orofacial clefts. Furthermore, NCC differentiation was impaired. Thus, NCC derivatives, such as skeletons, vasculatures and neural tissues were either rudimentary or malformed. We further demonstrate that disruption of mTOR caused P53 hyperactivity and cell cycle arrest in cranial NCCs, and lowering P53 activity by one copy reduction attenuated the severity of craniofacial phenotype in NCC-mTOR knockout mice. Remarkably, NCC-Rptor disruption caused a spectrum of defects mirroring that of the NCC-mTOR deletion, whereas NCC-Rictor disruption only caused a mild craniofacial phenotype compared to the mTOR and Rptor conditional knockout models. Altogether, our data demonstrate that mTOR functions mediated by mTORC1 are indispensable for multiple processes of NCC development including proliferation, survival, and differentiation during craniofacial morphogenesis and organogenesis, and P53 hyperactivity in part accounts for the defective craniofacial development in NCC-mTOR knockout mice.
Despite intensive effort was made to regenerate injured meniscus by cell-free strategies through recruiting endogenous stem/progenitor cells, meniscus regeneration remains a great challenge in ...clinic. In this study, we found decellularized meniscal extracellular matrix (MECM) preserved native meniscal collagen and glycosaminoglycans which could be a good endogenous regeneration guider for stem cells. Moreover, MECM significantly promoted meniscal fibrochondrocytes viability and proliferation, increased the expression of type II collagen and proteoglycans in vitro. Meanwhile, we designed 3D-printed polycaprolactone (PCL) scaffolds which mimic the circumferential and radial collagen orientation in native meniscus. Taken these two advantages together, a micro-structure and micro-environment dually biomimetic cell-free scaffold was manipulated. This cell-free PCL-MECM scaffold displayed superior biocompatibility and yielded favorable biomechanical capacities closely to native meniscus. Strikingly, neo-menisci were regenerated within PCL-MECM scaffolds which were transplanted into knee joints underwent medial meniscectomy in rabbits and sheep models. Histological staining confirmed neo-menisci showed meniscus-like heterogeneous staining. Mankin scores showed PCL-MECM scaffold could protect articular cartilage well, and knee X-ray examination revealed same results. Knee magnetic resonance imaging (MRI) scanning also showed some neo-menisci in PCL-MECM scaffold group. In conclusion, PCL-MECM scaffold appears to optimize meniscus regeneration. This could represent a promising approach worthy of further investigation in preclinical applications.
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•3D-printed PCL scaffolds could mimic the circumferential and radial collagen orientation in native meniscus.•PCL-MECM scaffold displayed superior biocompatibility and yielded favorable biomechanical capacities.•PCL-MECM scaffold appears to optimize meniscus regeneration in both rabbit and sheep meniscus repairing model.•PCL-MECM scaffold may represent a promising approach worthy of further investigation in preclinical applications.
Clonal reproduction (i.e., production of potentially independent offspring by vegetative growth) is thought to provide plants with reproductive assurance. Thus, studying the evolution of clonal ...reproduction in local floras is crucial for our understanding of the adaptive mechanisms plants deploy in stressful environments such as alpine regions. In this study, we characterized clonal plant species in the subnival belt of the Hengduan Mountains (a global biodiversity hotspot with extreme environmental conditions in southwest China), in order to determine the effects of sex system, growth form, and elevational distribution on clonality. We compiled clonality data of angiosperm species belonging to 41 families in the subnival belt of the Hengduan Mountains using published information. Of the 793 species recorded in the region, 47.92% (380 species) are clonal species. Both sex system and growth form had significant effects on the occurrence of clonal reproduction: unisexual species (79.79%) were more likely to be clonal than bisexual species (43.63%), and herbaceous species (51.04%) were more likely to be clonal than woody species (16.67%). Compared with non-alpine-endemic species (44.60%), alpine-endemic species (58.33%) showed a significantly higher proportion of clonal reproduction. Further logistic regression analysis showed a positive association between incidence of clonality and elevational range, indicating that species distributed at high elevations are more likely to be clonal. Furthermore, the elevational gradients in clonality were contingent on sex system or growth form. This study reveals that plants in the subnival belt of the Hengduan Mountains might optimize their probability of reproduction through clonal reproduction, a finding that adds to our growing understanding of plant's adaptations to harsh alpine environments.
To establish the three-dimensional facial soft tissue morphology of adolescent and adult females in the Guangdong population and to study the morphological characteristics of hyperdivergent skeletal ...class II females in Guangdong compared with that of normodivergent class I groups.
The 3dMDface system was used to capture face scans of 160 patients, including 45 normal and 35 hyperdivergent skeletal class II adolescents (aged 11-14 years old) and 45 normal and 35 hyperdivergent skeletal class II adults (aged 18-30 years old). Thirty-two soft tissue landmarks were mapped, and 21 linear, 10 angular and 17 ratio measurements were obtained by 3dMDvultus analysis software. Data were assessed with a t-test of two independent samples between the normal adolescent and adult groups and between the normal and hyperdivergent skeletal class II groups.
The linear measurements of the Guangdong adult females were larger than those of the adolescents in both Class I and Class II groups. However, the angular and ratio measurements had no significant difference. The vertical linear measurements were higher and the sagittal and transverse linear measurements were smaller in the hyperdivergent class II group (p < 0.05). The soft tissue ANB angle, chin-lip angle, and mandibular angle were significantly larger and the soft tissue facial convexity angle and nasal convexity angle were significantly smaller in the hyperdivergent class II group (p < 0.05). Additionally, there were significant differences in the ratio measurements between the hyperdivergent class II groups and the control groups (p < 0.05).
The three-dimensional facial morphology of Guangdong adolescent and adult females was acquired. The facial soft tissue measurements of the adults were higher in the three dimensions except for the facial convexity and proportional relationships which were similar, suggesting that the growth pattern remained the same. The three-dimensional facial soft tissue features of hyperdivergent skeletal class II were characterized by the terms "long, convex, and narrow". Three-dimensional facial measurements can reflect intrinsic hard tissue characteristics.
Periodontitis is a highly prevalent disease characterized by inflammation and destruction of tooth-supporting tissues that leads to tooth loss in extreme situations. Elucidating the underlying ...mechanisms of periodontitis pathogenesis and progression will establish the groundwork for developing effective treatment strategies. Recently, evidence concerning the role of ferroptosis in periodontitis progression has emerged. Osteogenic lineage cells are key regulators of bone remodeling. Osteogenic cell death, as observed in experimental periodontitis models, disrupts the balance between bone resorption and bone formation. However, whether the osteogenic lineage undergoes ferroptosis during periodontitis and the corresponding effect on periodontitis progression remain elusive. Here, we investigated cell-specific ferroptosis within the alveolar bone in a murine periodontitis model. Through immunofluorescence double staining and immunohistochemistry, we identified ferroptotic osteocytes and osteoblasts in inflammatory alveolar bone. Next, in vivo administration of erastin or liproxstatin-1 was conducted to either induce or inhibit ferroptosis, respectively. Severe bone resorption and inflammation, accompanied by increased osteoclast formation and impaired osteogenic potential were detected following ferroptosis activation. Subsequently, we carried out in vitro experiments on osteocytes and further verified that ferroptosis enhanced the osteocytic expression of RANKL and IL-6. These findings suggest that ferroptosis occurring within the osteogenic lineage acts as a catalyst in the progression of periodontitis by stimulating osteoclastogenesis through the secretion of inflammatory cytokines and inhibiting osteoblastic function, providing insights into ferroptosis-induced alterations in microenvironment-based intercellular communication. Ferroptosis is a promising target for controlling inflammation and preventing bone resorption in periodontitis.
Developmental defects of enamel are common due to genetic and environmental factors before and after birth. Cdc42, a Rho family small GTPase, regulates prenatal tooth development in mice. However, ...its role in postnatal tooth development, especially enamel formation, remains elusive. Here, we investigated Cdc42 functions in mouse enamel development and tooth repair after birth. Cdc42 showed highly dynamic temporospatial patterns in the developing incisors, with robust expression in ameloblast and odontoblast layers. Strikingly, epithelium-specific Cdc42 deletion resulted in enamel defects in incisors. Ameloblast differentiation was inhibited, and hypomineralization of enamel was observed upon epithelial Cdc42 deletion. Proteomic analysis showed that abnormal mitochondrial components, phosphotransferase activity, and ion channel regulator activity occurred in the Cdc42 mutant dental epithelium. Reactive oxygen species accumulation was detected in the mutant mice, suggesting that abnormal oxidative stress occurred after Cdc42 depletion. Moreover, Cdc42 mutant mice showed delayed tooth repair and generated less calcified enamel. Mitochondrial dysfunction and abnormal oxygen consumption were evidenced by reduced Apool and Timm8a1 expression, increased Atp5j2 levels, and reactive oxygen species overproduction in the mutant repair epithelium. Epithelium-specific Cdc42 deletion attenuated ERK1/2 signaling in the labial cervical loop. Aberrant Sox2 expression in the mutant labial cervical loop after clipping might lead to delayed tooth repair. These findings suggested that mitochondrial dysfunction, up-regulated oxidative stress, and abnormal ion channel activity may be among multiple factors responsible for the observed enamel defects in Cdc42 mutant incisors. Overall, Cdc42 exerts multidimensional and pivotal roles in enamel development and is particularly required for ameloblast differentiation and enamel matrix formation.
Osteoarthritis (OA) is a degenerative joint disease that is common among the middle-aged and older populations, causes patients to experience recurrent pain in their joints and negatively affects ...their quality of life. Currently, therapeutic options for patients with OA consist of medications to alleviate pain and treat the symptoms; however, due to typically poor outcomes, patients with advanced OA are unlikely to avoid joint replacement. In recent years, several studies have linked disrupted homeostasis of the joint cavity microenvironment to the development of OA. Recently, extracellular vesicles (EVs) have received increasing attention in the field of OA. EVs are natural nano-microcarrier materials with unique biological activity that are produced by cells through paracrine action. They are composed of lipid bilayers that contain physiologically active molecules, such as nucleic acids and proteins. Moreover, EVs may participate in local and distal intercellular and intracellular communication. EVs have also recently been shown to influence OA development by regulating biochemical factors in the OA microenvironmental. In this article, we first describe the microenvironment of OA. Then, we provide an overview of EVs, summarize the main types used for the treatment of OA, and describe their mechanisms. Next, we review clinical studies using EVs for OA treatment. Finally, the specific mechanism underlying the application of miRNA-enriched EVs in OA therapy is described.
Utilizing transplanted human umbilical cord mesenchymal stem cells (HUMSCs) for cartilage defects yielded advanced tissue regeneration, but the underlying mechanism remain elucidated. Early after ...HUMSCs delivery to the defects, we observed substantial apoptosis. The released apoptotic vesicles (apoVs) of HUMSCs promoted cartilage regeneration by alleviating the chondro-immune microenvironment. ApoVs triggered M2 polarization in macrophages while simultaneously facilitating the chondrogenic differentiation of endogenous MSCs. Mechanistically, in macrophages, miR-100-5p delivered by apoVs activated the MAPK/ERK signaling pathway to promote M2 polarization. In MSCs, let-7i-5p delivered by apoVs promoted chondrogenic differentiation by targeting the eEF2K/p38 MAPK axis. Consequently, a cell-free cartilage regeneration strategy using apoVs combined with a decellularized cartilage extracellular matrix (DCM) scaffold effectively promoted the regeneration of osteochondral defects. Overall, new mechanisms of cartilage regeneration by transplanted MSCs were unconcealed in this study. Moreover, we provided a novel experimental basis for cell-free tissue engineering-based cartilage regeneration utilizing apoVs.
Schematic summary of our main findings. Intraarticular injection of HUMSCs derived apoEVs delivers the miR-100-5p into macrophages and promotes M2 macrophage polarization by activating MAPK/ERK1/2 and delivers the let-7i-5p into MSCs and promotes chondrogenesis. The apoEVs combined with DCM scaffold facilitating cartilage defect repair. HUMSCs: human umbilical cord-derived mesenchymal stem cells, DCM: decellularized cartilage matrix. Display omitted
•This study provides the first evidence of short-term apoptosis of MSCs after implantion in cartilage defect.•HUMSC-apoVs induce macrophage immunomodulation, promoting MSC chondrogenic differentiation.•TmiR-100-5p in apoVs activated the MAPK/ERK pathway to induce macrophage M2 polarization.•Let-7i-5p in apoVs promoted chondrogenic differentiation by targeting the eEF2K/p38 MAPK axis.•A cell-free strategy of apoVs combined with DCM scaffold promoted cartilage regeneration.
Large bone defect reconstruction undergoes hypoxia and remains a major practical challenge. Bone tissue engineering with a more promising stem cell source facilitates the development of better ...therapeutic outcomes. Human dental follicle stem cells (hDFSCs) with superior multipotency, osteogenic capacity, and accessibility have been proven a promising cell source for bone regeneration. We previously identified a novel long noncoding RNA (lncRNA), HOTAIRM1, to be highly expressed in hDFSCs. Here we found that HOTAIRM1 overexpressed hDFSCs promoted bone regeneration in rat critical‐size calvarial defect model. Mechanically, HOTAIRM1 was induced in hDFSCs under hypoxic conditions and activated HIF‐1α. RNA‐sequencing analysis indicated that HOTAIRM1 upregulated oxygen‐sensing histone demethylases KDM6A/B and suppressed methyltransferase EZH2 via targeting HIF‐1α. The osteogenic differentiation of hDFSCs was accompanied with demethylation of H3K27, and HOTAIRM1 overexpression decreased the distribution of H3K27me3 in osteogenic genes, including ALP, M‐CSF, Wnt‐3a, Wnt‐5a, Wnt‐7a, and β‐catenin, thus promoted their transcription. Our study provided evidence that HOTAIRM1 upregulated KDM6A/B and inhibited EZH2 in a HIF‐1α dependent manner to enhance the osteogenesis of hDFSCs. HOTAIRM1‐mediated hDFSCs may serve as a promising therapeutic approach to promote bone regeneration in clinical practice.
HOTAIRM1 upregulated KDM6A/B and inhibited EZH2 in a HIF‐1α dependent manner to enhance the osteogenic differentiation of hDFSCs.
Organ development requires complex signaling by cells in different tissues. Epithelium and mesenchyme interactions are crucial for the development of skin, hair follicles, kidney, lungs, prostate, ...major glands, and teeth. Despite myriad literature on cell–cell interactions and ligand–receptor binding, the roles of extracellular vesicles in epithelium–mesenchyme interactions during organogenesis are poorly understood. Here, we discovered that ∼100 nm exosomes were secreted by the epithelium and mesenchyme of a developing tooth organ and diffused through the basement membrane. Exosomes were entocytosed by epithelium or mesenchyme cells with preference by reciprocal cells rather than self-uptake. Exosomes reciprocally evoked cell differentiation and matrix synthesis: epithelium exosomes induce mesenchyme cells to produce dentin sialoprotein and undergo mineralization, whereas mesenchyme exosomes induce epithelium cells to produce basement membrane components, ameloblastin and amelogenenin. Attenuated exosomal secretion by Rab27a/b knockdown or GW4869 disrupted the basement membrane and reduced enamel and dentin production in organ culture and reduced matrix synthesis and the size of the cervical loop, which harbors epithelium stem cells, in Rab27aash/ash mutant mice. We then profiled exosomal constituents including miRNAs and peptides and further crossed all epithelium exosomal miRNAs with literature-known miRNA Wnt regulators. Epithelium exosome-derived miR135a activated Wnt/β-catenin signaling and escalated mesenchymal production of dentin matrix proteins, partially reversible by Antago-miR135a attenuation. Our results suggest that exosomes may mediate epithelium–mesenchyme crosstalk in organ development, suggesting that these vesicles and/or the molecular contents they are transporting may be interventional targets for treatment of diseases or regeneration of tissues.
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