RESUMO
Radiation therapy is a clinically proven, localized preventive measure for heterotopic ossification (HO). Despite its efficacy, there is a lack of standardization of radiation prescription dosing and fractionation, and the mechanism of the impact of radiation in HO prevention remains unknown. Here, using an established mouse model of traumatic HO induced by burn and tenotomy, we demonstrate that 7Gy in one fraction delivered to the injury site within 72 hours postoperatively significantly decreases HO formation and improves hindlimb range of motion. In-depth single-cell transcriptomic analyses, in combination with immunofluorescent staining, demonstrate decreased cellular numbers as well as aberrant endochondral differentiation and downregulation of associated upstream signaling pathways in irradiated mesenchymal progenitor cells. Our study provides the framework for future mechanistic and clinically relevant studies exploring radiation efficacy in preventing HO formation.
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While hypoxic signaling has been shown to play a role in many cellular processes, its role in metabolism-linked extracellular matrix (ECM) organization and downstream processes of cell fate after musculoskeletal injury remains to be determined. Heterotopic ossification (HO) is a debilitating condition where abnormal bone formation occurs within extra-skeletal tissues. Hypoxia and hypoxia-inducible factor 1α (HIF-1α) activation have been shown to promote HO. However, the underlying molecular mechanisms by which the HIF-1α pathway in mesenchymal progenitor cells (MPCs) contributes to pathologic bone formation remain to be elucidated. Here, we used a proven mouse injury-induced HO model to investigate the role of HIF-1α on aberrant cell fate. Using single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics analyses of the HO site, we found that collagen ECM organization is the most highly up-regulated biological process in MPCs. Zeugopod mesenchymal cell-specific deletion of Hif1α (Hoxa11-CreERT2; Hif1afl/fl) significantly mitigated HO in vivo. ScRNA-seq analysis of these Hoxa11-CreERT2; Hif1afl/fl mice identified the PLOD2/LOX pathway for collagen cross-linking as downstream of the HIF-1α regulation of HO. Importantly, our scRNA-seq data and mechanistic studies further uncovered that glucose metabolism in MPCs is most highly impacted by HIF-1α deletion. From a translational aspect, a pan-LOX inhibitor significantly decreased HO. A newly screened compound revealed that the inhibition of PLOD2 activity in MPCs significantly decreased osteogenic differentiation and glycolytic metabolism. This suggests that the HIF-1α/PLOD2/LOX axis linked to metabolism regulates HO-forming MPC fate. These results suggest that the HIF-1α/PLOD2/LOX pathway represents a promising strategy to mitigate HO formation.
Assuntos
Ossificação Heterotópica , Osteogênese , Animais , Camundongos , Colágeno/metabolismo , Modelos Animais de Doenças , Matriz Extracelular/genética , Hipóxia/metabolismo , Ossificação Heterotópica/metabolismo , Fatores de Transcrição/metabolismoRESUMO
Heterotopic ossification (HO) is a debilitating pathology where ectopic bone develops in areas of soft tissue. HO can develop as a consequence of traumatic insult or as a result of dysregulated osteogenic signaling, as in the case of the orphan disease fibrodysplasia ossificans progressiva (FOP). Traumatic HO (tHO) formation is mediated by the complex interplay of signaling between progenitor, inflammatory, and nerve cells, among others, making it a challenging process to understand. Research into the pathogenesis of genetically mediated HO (gHO) in FOP has established a pathway involving uninhibited activin-like kinase 2 receptor (ALK2) signaling that leads to downstream osteogenesis. Current methods of diagnosis and treatment lag behind pre-mature HO detection and progressive HO accumulation, resulting in irreversible decreases in range of motion and chronic pain for patients. As such, it is necessary to draw on advancements made in the study of tHO and gHO to better diagnose, comprehend, prevent, and treat both.
Assuntos
Miosite Ossificante , Ossificação Heterotópica , Humanos , Miosite Ossificante/diagnóstico , Miosite Ossificante/genética , Miosite Ossificante/complicações , Ossificação Heterotópica/etiologia , Ossificação Heterotópica/metabolismo , Ossificação Heterotópica/patologia , Osteogênese , Osso e Ossos/metabolismoRESUMO
Hypothalamic Kiss1 neurons control gonadotropin-releasing hormone release through the secretion of kisspeptin. Kiss1 neurons serve as a nodal center that conveys essential regulatory cues for the attainment and maintenance of reproductive function. Despite this critical role, the mechanisms that control kisspeptin synthesis and release remain largely unknown. Using Drop-Seq data from the arcuate nucleus of adult mice and in situ hybridization, we identified Nescient Helix-Loop-Helix 2 (Nhlh2), a transcription factor of the basic helix-loop-helix family, to be enriched in Kiss1 neurons. JASPAR analysis revealed several binding sites for NHLH2 in the Kiss1 and Tac2 (neurokinin B) 5' regulatory regions. In vitro luciferase assays evidenced a robust stimulatory action of NHLH2 on human KISS1 and TAC3 promoters. The recruitment of NHLH2 to the KISS1 and TAC3 promoters was further confirmed through chromatin immunoprecipitation. In vivo conditional ablation of Nhlh2 from Kiss1 neurons using Kiss1Cre:Nhlh2fl/fl mice induced a male-specific delay in puberty onset, in line with a decrease in arcuate Kiss1 expression. Females retained normal reproductive function albeit with irregular estrous cycles. Further analysis of male Kiss1Cre:Nhlh2fl/fl mice revealed higher susceptibility to metabolic challenges in the release of luteinizing hormone and impaired response to leptin. Overall, in Kiss1 neurons, Nhlh2 contributes to the metabolic regulation of kisspeptin and NKB synthesis and release, with implications for the timing of puberty onset and regulation of fertility in male mice.
Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/fisiologia , Kisspeptinas/metabolismo , Neurônios/fisiologia , Maturidade Sexual/fisiologia , Animais , Linhagem Celular , Cromatina , DNA/genética , Estradiol/farmacologia , Feminino , Fertilidade , Regulação da Expressão Gênica/efeitos dos fármacos , Imunoprecipitação , Kisspeptinas/genética , Kisspeptinas/farmacologia , Leptina/farmacologia , Hormônio Luteinizante/metabolismo , Masculino , Camundongos , Camundongos Knockout , Fragmentos de Peptídeos/farmacologia , Reação em Cadeia da Polimerase/métodos , Fatores Sexuais , Substância P/análogos & derivados , Substância P/farmacologiaRESUMO
The alternation of the stimulatory action of the tachykinin neurokinin B (NKB) and the inhibitory action of dynorphin within arcuate (ARH) Kiss1 neurons has been proposed as the mechanism behind the generation of gonadotropin-releasing hormone (GnRH) pulses through the pulsatile release of kisspeptin. However, we have recently documented that GnRH pulses still exist in gonadectomized mice in the absence of tachykinin signaling. Here, we document an increase in basal frequency and amplitude of luteinizing hormone (LH) pulses in intact male mice deficient in substance P, neurokinin A (NKA) signaling (Tac1KO), and NKB signaling (Tac2KO and Tacr3KO). Moreover, we offer evidence that a single bolus of the NKB receptor agonist senktide to gonad-intact wild-type males increases the basal release of LH without changing its frequency. Altogether, these data support the dispensable role of the individual tachykinin systems in the generation of LH pulses. Moreover, the increased activity of the GnRH pulse generator in intact KO male mice suggests the existence of compensation by additional mechanisms in the generation of kisspeptin/GnRH pulses.
Assuntos
Hormônio Luteinizante/sangue , Receptores da Neurocinina-3/metabolismo , Taquicininas/metabolismo , Animais , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores da Neurocinina-3/genética , Taquicininas/genéticaRESUMO
The d1T-MoS2, distorted-1T group-VIB transition metal dichalcogenides monolayer, is considered as promising atomically thin out-of-plane ferroelectric materials. We study the origin of the ferroelectricity in d1T-MoS2 monolayer using first-principles calculations and the Landau theory of phase transition. In contrast to conventional improper ferroelectrics, we find that the polarization has dependence on both primary and secondary modes. It turns out that the charge imbalance between chalcogen atoms at different symmetry sites is the source for the out-of-plane polarization without any out-of-plane displacement in the primary mode. The secondary mode following the primary mode cancels partially the polarization and secures it against the depolarization field. We show that the polarization of d1T-MoS2 is robust to external electric fields and can be manipulated by the uniaxial strain.
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Novel probe development for positron emission tomography (PET) is leading to expanding the scope of molecular imaging. To begin responding to challenges, several biomaterials such as natural products and small molecules, peptides, engineered proteins including affibodies, and antibodies have been used in the development of targeted molecular imaging probes. To prepare radiotracers, a few bioactive materials are unique challenges to radiolabelling because of their complex structure, poor stability, poor solubility in aqueous or chemical organic solutions, and sensitivity to temperature and nonphysiological pH. To overcome these challenges, we developed a new radiolabelling strategy based on photoactivated 1,3-dipolar cycloaddition between alkene dipolarophile and tetrazole moiety containing compounds. Herein, we describe a light-triggered radiochemical synthesis via photoactivated click reaction to prepare 18F-radiolabelled PET tracers using small molecular and RGD peptide.
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Química Click/métodos , Marcação por Isótopo/métodos , Tomografia por Emissão de Pósitrons/métodos , Compostos Radiofarmacêuticos/síntese química , Alcenos/química , Animais , Linhagem Celular Tumoral , Reação de Cicloadição , Radioisótopos de Flúor/química , Xenoenxertos , Humanos , Luz , Sondas Moleculares/síntese química , Ratos , Tetrazóis/químicaRESUMO
Group-VIIB transition metal dichalcogenides (TMDCs) are known to be stabilized solely in a distorted 1T phase termed as 1Tâ³ phase, which is compared to many stable or metastable phases in other TMDCs. Using first-principles calculations, we study the structural origin of 1Tâ³ phase group-VIIB TMDCs. We find that quasi 1D Peierls-like instability is responsible for the transition to the 1Tâ³ phase ReS2 monolayer from the 1T' phase, another distorted 1T phase. Two half-filled bands in 1T'-ReS2 make sharp peaks in the Lindhard function that prompt the charge density wave (CDW) phase with large band gap opening. Our calculations show that overlapping of the two bands in a broad energy range leads to robust CDW phase or stable 1Tâ³ phase in group-VIIB TMDCs against compositional variation, which is in stark contrast to typical Peierls instability driven by a single band. Calculated total energy curve near the critical point exhibits the feature of the first-order Landau transition due to local chemical bonding. The structural stability of the 1Tâ³ phase in group-VIIB TMDCs is thus guaranteed by two half-filled bands and local chemical bonding.
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Methicillinresistant Staphylococcus aureus (MRSA) is difficult to treat using available antibiotic agents. Honeybee venom has been widely used as an oriental treatment for several inflammatory diseases and bacterial infections. The venom contains predominantly biologically active compounds, however, the therapeutic effects of such materials when used to treat MRSA infections have not been investigated extensively. The present study evaluated bee venom and its principal active component, melittin, in terms of their antibacterial activities and in vivo protection against MRSA infections. In vitro, bee venom and melittin exhibited comparable levels of antibacterial activity, which was more marked against MRSA strains, compared with other Grampositive bacteria. When MRSAinfected mice were treated with bee venom or melittin, only the latter animals were successfully rescued from MRSA induced bacteraemia or exhibited recovery from MRSAinfected skin wounds. Together, the data of the present study demonstrated for the first time, to the best of our knowledge, that melittin may be used as a promising antimicrobial agent to enhance the healing of MRSAinduced wounds.
Assuntos
Antibacterianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Venenos de Abelha/química , Meliteno/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Infecções Estafilocócicas/tratamento farmacológico , Animais , Antibacterianos/síntese química , Antibacterianos/isolamento & purificação , Peptídeos Catiônicos Antimicrobianos/síntese química , Peptídeos Catiônicos Antimicrobianos/isolamento & purificação , Abelhas/química , Abelhas/fisiologia , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Humanos , Células MCF-7 , Masculino , Meliteno/síntese química , Meliteno/isolamento & purificação , Staphylococcus aureus Resistente à Meticilina/crescimento & desenvolvimento , Staphylococcus aureus Resistente à Meticilina/isolamento & purificação , Camundongos , Testes de Sensibilidade Microbiana , Infecções Estafilocócicas/microbiologia , Infecções Estafilocócicas/mortalidade , Infecções Estreptocócicas/microbiologia , Streptococcus/efeitos dos fármacos , Streptococcus/crescimento & desenvolvimento , Streptococcus/isolamento & purificação , Análise de SobrevidaRESUMO
Staphylococcus aureus (S. aureus) is a virulent bacterium that abundantly colonizes inflammatory skin diseases. Since S. aureus infections occur in an impaired skin barrier, it is important to understand the protective mechanism through cutaneous immune responses against S. aureus infections and the interaction with Staphylococcal virulence factors. In this review, we summarize not only the pathogenesis and key elements of S. aureus skin infections, but also the cutaneous immune system against its infections and colonization. The information obtained from this area may provide the groundwork for further immunomodulatory therapies or vaccination strategies to prevent S. aureus infections.
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We investigated whether sulodexide has additional protective effects against peripheral nerve damage caused by microvascular dysfunction in a rat model of diabetes. Female Sprague-Dawley (SD) rats were divided into the following 4 groups (n=7-9/group): Normal, Normal+Sulodexide (sulodexide 10mg/kg), diabetic group, and diabetic+Sulodexide (sulodexide 10mg/kg). We assessed current perception threshold, skin blood flow, superoxide dismutase, and proteinuria in experimental rats after oral administration of sulodexide for 20 weeks. We also performed morphometric analysis of sciatic nerves and intraepidermal nerve fibers of the foot. Superoxide dismutase activity in the blood and sciatic nerve were increased significantly after sulodexide treatment in the diabetic group. Current perception threshold was reduced at 2000 Hz (633.3 ± 24.15 vs 741.2 ± 23.5 µA, P<0.05) and skin blood flow was improved (10.90 ± 0.67 vs 8.85 ± 0.49 TPU, P<0.05) in the diabetic+Sulodexide group compared with the diabetic group. The mean myelinated axon area was significantly larger (56.6 ± 2.2 vs 49.8 ± 2.7 µm(2), P<0.05) and the intraepidermal nerve fiber density was significantly less reduced (6.27 ± 0.24 vs 5.40 ± 0.25/mm, P<0.05) in the diabetic+Sulodexide group compared to the diabetic group. Our results demonstrate that sulodexide exhibits protective effects against peripheral nerve damage in a rat experimental model of diabetes. Therefore, these findings suggest that sulodexide is a potential new therapeutic agent for diabetic peripheral neuropathy.