RESUMO
BACKGROUND: Tick bites severely threaten human health because they allow the transmission of many deadly pathogens, including viruses, bacteria, protozoa, and helminths. Pruritus is a leading symptom of tick bites, but its molecular and neural bases remain elusive. OBJECTIVES: This study sought to discover potent drugs and targets for the specific prevention and treatment of tick bite-induced pruritus and arthropod-related itch. METHODS: We used live-cell calcium imaging, patch-clamp recordings, and genetic ablation and evaluated mouse behavior to investigate the molecular and neural bases of tick bite-induced pruritus. RESULTS: We found that 2 tick salivary peptides, IP defensin 1 (IPDef1) and IR defensin 2 (IRDef2), induced itch in mice. IPDef1 was further revealed to have a stronger pruritogenic potential than IRDef2 and to induce pruritus in a histamine-independent manner. IPDef1 evoked itch by activating mouse MrgprC11 and human MRGPRX1 on dorsal root ganglion neurons. IPDef1-activated MrgprC11/X1 signaling sensitized downstream ion channel TRPV1 on dorsal root ganglion neurons. Moreover, IPDef1 also activated mouse MrgprB2 and its ortholog human MRGPRX2 selectively expressed on mast cells, inducing the release of inflammatory cytokines and driving acute inflammation in mice, although mast cell activation did not contribute to oxidated IPDef1-induced itch. CONCLUSIONS: Our study identifies tick salivary peptides as a new class of pruritogens that initiate itch through MrgprC11/X1-TRPV1 signaling in pruritoceptors. Our work will provide potential drug targets for the prevention and treatment of pruritus induced by the bites or stings of tick and maybe other arthropods.
Assuntos
Peptídeos/imunologia , Receptores Acoplados a Proteínas G/metabolismo , Canais de Cátion TRPV/metabolismo , Carrapatos/imunologia , Alérgenos/imunologia , Animais , Suscetibilidade a Doenças , Humanos , Camundongos , Prurido/imunologia , Prurido/metabolismoRESUMO
Viral infections still threaten human health all over the world, and many people die from viral diseases every year. However, there are no effective vaccines or drugs for preventing or managing most viral diseases. Thus, the discovery and development of broad-spectrum antiviral agents remain urgent. Here, we expressed and purified a venom peptide, Ev37, from the scorpion Euscorpiops validus in a prokaryotic system. We found that rEv37 can inhibit dengue virus type 2 (DENV-2), hepatitis C virus (HCV), Zika virus (ZIKV), and herpes simplex virus type 1 (HSV-1) infections in a dose-dependent manner at noncytotoxic concentrations, but that it has no effect on Sendai virus (SeV) and adenovirus (AdV) infections in vitro Furthermore, rEv37 alkalized acidic organelles to prevent low pH-dependent fusion of the viral membrane-endosomal membrane, which mainly blocks the release of the viral genome from the endosome to the cytoplasm and then restricts viral late entry. Taken together, our results indicate that the scorpion venom peptide Ev37 is a broad-spectrum antiviral agent with a specific molecular mechanism against viruses undergoing low pH-dependent fusion activation during entry into host cells. We conclude that Ev37 is a potential candidate for development as an antiviral drug.
Assuntos
Citoplasma/metabolismo , Vírus da Dengue/fisiologia , Endossomos/metabolismo , Venenos de Escorpião/farmacologia , Escorpiões/química , Internalização do Vírus/efeitos dos fármacos , Adenoviridae/fisiologia , Animais , Chlorocebus aethiops , Citoplasma/virologia , Endossomos/virologia , Células HEK293 , Humanos , Fusão de Membrana/efeitos dos fármacos , Venenos de Escorpião/química , Vírus Sendai/fisiologia , Células VeroRESUMO
The outbreak of coronavirus disease 2019 (COVID-19) has spread across the world and was characterized as a pandemic. To protect medical laboratory personnel from infection, most laboratories inactivate the virus causing COVID-19, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), in clinical samples before testing. However, the effect of inactivation on the detection results remains unknown. Here, we used a digital PCR assay to determine the absolute SARS-CoV-2 RNA copy number in 63 nasopharyngeal swab samples and assess the effect of inactivation methods on viral RNA copy number. Viral inactivation was performed by three different methods: (i) incubation with the TRIzol LS reagent for 10 min at room temperature, (ii) heating in a water bath at 56°C for 30 min, and (iii) high-temperature treatment, including autoclaving at 121°C for 20 min, boiling at 100°C for 20 min, and heating at 80°C for 20 min. Compared to the amount of RNA in the original sample, TRIzol treatment destroyed 47.54% of the nucleocapsid protein (N) gene and 39.85% of open reading frame (ORF) 1ab. For samples treated at 56°C for 30 min, the copy number of the N gene and ORF 1ab was reduced by 48.55% and 56.40%, respectively. The viral RNA copy number dropped by 50 to 66% after heating at 80°C for 20 min. Nearly no viral RNA was detected after autoclaving at 121°C or boiling at 100°C for 20 min. These results indicate that inactivation reduced the quantity of detectable viral RNA and may cause false-negative results, especially in weakly positive cases. Thus, use of the TRIzol reagent rather than heat inactivation is recommended for sample inactivation, as the TRIzol reagent had the least effect on the RNA copy number among the tested methods.
Assuntos
Betacoronavirus/efeitos dos fármacos , Betacoronavirus/efeitos da radiação , Desinfecção/métodos , RNA Viral/análise , Manejo de Espécimes/métodos , Inativação de Vírus/efeitos dos fármacos , Inativação de Vírus/efeitos da radiação , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Desinfetantes , Feminino , Dosagem de Genes , Temperatura Alta , Humanos , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , RNA Viral/genética , SARS-CoV-2 , Adulto JovemRESUMO
BACKGROUNDS: Abdominal obesity has been linked to risk of mortality, but whether and how trajectory of waist circumstance (WC) underpins this association remains unclear. The study aimed to identify long-term WC change trajectories and examine their association and joint effect with body mass index (BMI) on mortality among Chinese older adults. METHODS: This present study included participants 60 years of age or older from China Health and Nutrition Survey (CHNS) from 1991 to 2015. The duration of follow-up was defined as period from the first to latest visit date attended with information on mortality, end of follow-up, or loss to follow-up (censoring). Latent class trajectory analysis (LCTA) was used to assess the changes of WC trajectories overtime. Cox proportional hazard models were used to assess hazard ratios (HRs) and corresponding 95% confidence internal (CIs) for mortality. RESULTS: A total of 2601 participants with 8700 visits were included, and 562 mortality (21.6%) occurred during a median follow-up of 8.7 years. Using a group-based modeling approach, four distinct trajectories of WC change among Chinese older adults were identified as loss (13.5%), stable (46.8%), moderate gain (31.2%) and substantial gain (8.5%). With WC stable group as reference, the multivariable adjusted HRs for mortality were 1.34(95%CI:1.01-1.78) in loss group, 1.13(0.91-1.41) in moderate gain and 1.54(1.12-2.12) in substantial gain group. Compared with participants with normal BMI at baseline and maintained WC stable, the risk of mortality generally increased for all WC change group in initial overweight/obesity individuals, and the highest risk were observed for WC loss and stable pattern (HR:2.43, 95%CI: 1.41-4.19; HR:1.67 (1.07-2.60)). CONCLUSIONS: In older Chinese, both long-term WC loss and substantial gain conferred excess risk for mortality. The baseline BMI might modify the effect as overweight individuals had a greater risk imposed by WC loss than those in normal weight. Maintaining stable WC and normal weight might be necessary to reduce the risk of mortality.
RESUMO
As a pathogenic microorganism, Listeria monocytogenes is widely used in the research of bacterial pathogenesis and host defense. The phagosomal escape of L. monocytogenes is essential for its replication in the cytoplasm of the host. Here, we reported that the protein abundance of the Six-transmembrane epithelial antigen of the prostate 3 (Steap3) was decreased upon L. monocytogenes infection compared to uninfected cells in macrophages. However, the decreased Steap3 abundance was not regulated by the host but was caused by LLO secreted by L. monocytogenes. Functional experiments showed that deletion of Steap3 facilitated entry of L. monocytogenes from the phagosome into the cytoplasm. Then, the comprehensive proteomic analysis revealed that the deletion of Steap3 could affect the proteins abundance of the lysosomal signaling pathway in macrophages. Among these proteins affected by Steap3, we discovered that only the Ganglioside GM2 activator (Gm2a) inhibited the phagosomal escape of L. monocytogenes as Steap3. In summary, we found that the Steap3-Gm2a axis could restrict the phagosomal escape of L. monocytogenes and serve the potential molecular drug targets for antibacterial treatment.
Assuntos
Toxinas Bacterianas , Listeria monocytogenes , Listeriose , Masculino , Humanos , Listeriose/microbiologia , Gangliosídeo G(M2)/metabolismo , Proteínas Hemolisinas/metabolismo , Gangliosídeos/metabolismo , Proteômica , Toxinas Bacterianas/metabolismo , Proteínas de Choque Térmico/metabolismo , Fagossomos/microbiologiaRESUMO
Interferon-inducible transmembrane proteins (IFITM1/2/3) have been reported to suppress the entry of a wide range of viruses. However, their antiviral functional residues and specific mechanisms are still unclear. Here, we firstly resolved the topology of IFITM1 on the plasma membrane where N-terminus points into the cytoplasm and C-terminus resides extracellularly. Further, KRRK basic residues of IFITM1 locating at 62-67 of the conserved intracellular loop (CIL) were found to play a key role in the restriction on the Zika virus (ZIKV) and dengue virus (DENV). Similarly, KRRK basic residues of IFITM2/3 also contributed to suppressing ZIKV replication. Finally, IFITM1 was revealed to be capable of restricting the release of ZIKV particles from endosome to cytosol so as to impede the entry of ZIKV into host cells, which was tightly related with the inhibition of IFITM1 on the acidification of organelles. Overall, our study provided topology, antiviral functional residues and the mechanism of interferon-inducible transmembrane proteins.
Assuntos
Antígenos de Diferenciação/metabolismo , Antígenos de Diferenciação/farmacologia , Antivirais/metabolismo , Antivirais/farmacologia , Zika virus/efeitos dos fármacos , Sequência de Aminoácidos , Animais , Membrana Celular/metabolismo , Chlorocebus aethiops , Endossomos/metabolismo , Células HEK293 , Humanos , Interferons/metabolismo , Proteínas de Membrana/metabolismo , Alinhamento de Sequência , Células Vero , Replicação Viral/efeitos dos fármacos , Infecção por Zika virusRESUMO
The objective of this study was to assess the effects of exogenously expressed proinsulin on the biological characters of a hematopoietic stem cell line (HSC) and erythroid myeloid lymphoid (EML) cells and explore new strategies for cell therapy for type I diabetes. EML cells were transduced with lentivirus particles carrying the human proinsulin (proINS) gene. The positive transduced cells were selected based on green fluorescence protein (GFP) positivity and puromycin resistance. Overexpression of proINS was confirmed via real-time PCR and Western blotting. The functional activity of the human proINS secreted by EML cells was elucidated by analyzing the activation of insulin receptor and its downstream signaling. Pro-INS + EML cells were able to prime the phosphorylation of insulin receptor as well as induce the expression of downstream genes of insulin receptor. Furthermore, Wnt3a can significantly promote self-renewal of Pro-INS + EML cells. However, we did not observe significant changes in the proliferation and differentiation of INS + EML cells, compared to the control EML cells. Our results might be useful for developing a new therapy for diabetes mellitus.
Assuntos
Diferenciação Celular , Proliferação de Células , Autorrenovação Celular , Células-Tronco Hematopoéticas/metabolismo , Proinsulina/biossíntese , Antígenos CD/genética , Antígenos CD/metabolismo , Linhagem Celular , Células Precursoras Eritroides/metabolismo , Humanos , Fosforilação , Proinsulina/genética , Receptor de Insulina/genética , Receptor de Insulina/metabolismo , Transdução de Sinais , Fatores de Tempo , Transdução Genética , Transfecção , Regulação para Cima , Proteína Wnt3A/metabolismoRESUMO
Epidemiological studies have validated the association between hepatitis C virus (HCV) infection and hepatocellular carcinoma (HCC). An increasing number of studies show that protein-protein interactions (PPIs) between HCV proteins and host proteins play a vital role in infection and mediate HCC progression. In this work, we collected all published interaction between HCV and human proteins, which include 455 unique human proteins participating in 524 HCV-human interactions. Then, we construct the HCV-human and HCV-HCC protein interaction networks, which display the biological knowledge regarding the mechanism of HCV pathogenesis, particularly with respect to pathogenesis of HCC. Through in-depth analysis of the HCV-HCC interaction network, we found that interactors are enriched in the JAK/STAT, p53, MAPK, TNF, Wnt, and cell cycle pathways. Using a random walk with restart algorithm, we predicted the importance of each protein in the HCV-HCC network and found that AKT1 may play a key role in the HCC progression. Moreover, we found that NS5A promotes HCC cells proliferation and metastasis by activating AKT/GSK3ß/ß-catenin pathway. This work provides a basis for a detailed map tracking new cellular interactions of HCV and identifying potential targets for HCV-related hepatocellular carcinoma treatment.