Alpha-synuclein dynamics bridge Type-I Interferon response and SARS-CoV-2 replication in peripheral cells

BACKGROUND: Increasing evidence suggests a double-faceted role of alpha-synuclein (α-syn) following infection by a variety of viruses, including SARS-CoV-2. Although α-syn accumulation is known to contribute to cell toxicity and the development and/or exacerbation of neuropathological manifestations, it is also a key to sustaining anti-viral innate immunity. Consistently with α-syn aggregation as a hallmark of Parkinson's disease, most studies investigating the biological function of α-syn focused on neural cells, while reports on the role of α-syn in periphery are limited, especially in SARS-CoV-2 infection. RESULTS: Results herein obtained by real time qPCR, immunofluorescence and western blot indicate that α-syn upregulation in peripheral cells occurs as a Type-I Interferon (IFN)-related response against SARS-CoV-2 infection. Noteworthy, this effect mostly involves α-syn multimers, and the dynamic α-syn multimer:monomer ratio. Administration of excess α-syn monomers promoted SARS-CoV-2 replication along with downregulation of IFN-Stimulated Genes (ISGs) in epithelial lung cells, which was associated with reduced α-syn multimers and α-syn multimer:monomer ratio. These effects were prevented by combined administration of IFN-ß, which hindered virus replication and upregulated ISGs, meanwhile increasing both α-syn multimers and α-syn multimer:monomer ratio in the absence of cell toxicity. Finally, in endothelial cells displaying abortive SARS-CoV-2 replication, α-syn multimers, and multimer:monomer ratio were not reduced following exposure to the virus and exogenous α-syn, suggesting that only productive viral infection impairs α-syn multimerization and multimer:monomer equilibrium. CONCLUSIONS: Our study provides novel insights into the biology of α-syn, showing that its dynamic conformations are implicated in the innate immune response against SARS-CoV-2 infection in peripheral cells. In particular, our results suggest that promotion of non-toxic α-syn multimers likely occurs as a Type-I IFN-related biological response which partakes in the suppression of viral replication. Further studies are needed to replicate our findings in neuronal cells as well as animal models, and to ascertain the nature of such α-syn conformations.

Construction of interferon alpha/beta receptor subunit 1 gene knockout Caco-2 cell line based on CRISPR/Cas9 system / 中国生物制品学杂志

@#Objective To knockout interferon alpha/beta receptor subunit 1(IFNAR1) gene in human colorectal adenocarcinoma cells Caco-2 using clustered regularly interspaced short palinmic repeats(CRISPR)/CRISPR-associated protein 9(Cas9)system to construct IFNAR1 knockout Caco-2 cell line.Methods The single guide RNA(sgRNA)sequence was designed to specifically recognize the exon region of IFNAR1 gene using CRISPR/Cas9 technology,and the LentiCRISPRv2-IFNAR1-sgRNA recombinant plasmid was constructed.Caco-2 cells were infected with the plasmid packaged by lentivirus and screened by puromycin resistance.The obtained monoclonal cell lines were cultured by limited dilution method,which were verified for the effect of IFNAR1 gene knockout by target gene sequencing and Western blot,and detected for the mRNA levels of CXC chemokine ligand 10(CXCL10)and interferon-stimulatd gene 20(ISG20)in IFNAR1knockout cells by adding exogenous IFNβ.Results Sequencing results of plasmid LentiCRISPRv2-IFNAR1-sgRNA showed that the insertion sites were all located at the sticky end of BsmBⅠenzyme digestion.Two IFNAR1 knockout monoclonal cell lines were obtained.The sequencing results showed that Caco-2-IFNAR1-KO1 had 5 bp deletion in the sixth exon of IFNAR1,and Caco-2-IFNAR1-KO2 had 18 bp deletion and 1 bp insertion in the seventh exon.Compared with wild-type Caco-2 cells,Caco-2-IFNAR1-KO1 and Caco-2-IFNAR1-KO2 cells showed no expression of IFNAR1 protein.Compared with no IFNβ stimulation,the mRNA levels of CXCL10 gene(t = 0.566 and 1.268 respectively,P>0.05)and ISG20 gene(t =1.522 and 1.733 respectively,P>0.05)in Caco-2-IFNAR1-KO1 and Caco-2-IFNAR1-KO2 cells stimulated by 50 ng/mL IFNβ showed no significant increase.While compared with those of wild-type Caco-2 cells,the mRNA levels of CXCL10gene(t = 6.763 and 6.777 respectively,P<0.05)and ISG20 gene(t = 5.664 and 5.65 respectively,P<0.05)in Caco-2-IFNAR1-KO1 and Caco-2-IFNAR1-KO2 cells decreased significantly under the stimulation of 50 ng/mL exogenous IFNβ.Conclusion Caco-2 cell line with IFNAR1 knockout was successfully constructed by using CRISPR/Cas9 technology,and the downstream molecules activated by IFNAR(interferon alpha/beta receptor)in this cell line were obviously inhibited,which provided a powerful tool for further exploration of the innate immune response and replication packaging mechanism of Caco-2 cells after virus infection.

Expressions of long non-coding RNA LINC00673 and ISG15 protein in pancreatic cancer and their clinical significances / 肿瘤研究与临床

Objective:To explore the expressions of long non-coding RNA LINC00673 and ISG15 protein in pancreatic cancer and their clinical significances.Methods:The clinical data of 57 patients diagnosed as pancreatic ductal carcinoma (PDAC) at the Affiliated Cancer Hospital of Xiangya Medical College of Central South University from January 2014 to December 2018 were retrospectively analyzed. The relative expressions of LINC00673 in pancreatic cancer tissues and paracancerous normal tissues (within 3 cm from the edge of cancer tissues) were examined by using quantificational reverse transcription-polymerase chain reaction (qRT-PCR). The ISG15 protein expressions in pancreatic cancer tissues and paracancerous normal tissues were examined by using immunohistochemistry. The difference in LINC00673 expression between ISG15 protein positive and negative patients was compared. The correlation between LINC00673 and ISG15 protein expressions in pancreatic cancer was analyzed by Spearman rank correlation analysis. Moreover, the correlations of LINC00673 and ISG15 protein expressions with clinical stage and pathological classification of pancreatic cancer patients were analyzed.Results:The positive expression of ISG15 protein in pancreatic cancer tissues was 40.4% (23/57), which was higher than that in paracancerous normal tissues [15.8% (9/57)] ( χ2 = 7.90, P = 0.004), and the relative expression of LINC00673 in pancreatic cancer tissues was 0.99±0.36, which was lower than that in paracancerous normal tissues (1.26±0.41) ( t = 4.80, P < 0.001). For 23 (40.4%) ISG15-positive patients and 34 (59.7%) ISG15-negative patients, the relative expression of LINC00673 was 0.77±0.46 and 0.45±0.27 ( P < 0.001). Spearman analysis showed that there was a correlation between LINC00673 and ISG15 protein expressions ( ρ = -0.429, P = 0.001). The relative expression of LINC00673 decreased in patients with low differentiated or undifferentiated tumor, vascular invasion and lymph node metastasis (all P < 0.05), but there was no correlation between LINC00673 expression and patients' age, tumor site, preoperative CA199 level, and TNM stage (all P > 0.05); ISG15 protein expression increased in patients with low differentiated or undifferentiated tumor, TNM stage Ⅲ-Ⅳ, vascular invasion and lymph node metastasis (all P < 0.05), but there was no correlation between ISG15 protein expression and patients' gender, age, tumor site, and preoperative CA199 level (all P > 0.05). Conclusions:The expression of LINC00673 in pancreatic cancer is related to vascular invasion, tumor differentiation degree and lymph node metastasis, and the expression of ISG15 in pancreatic cancer is related to vascular invasion, tumor differentiation degree, lymph node metastasis and TNM stage. The combined detection of LINC00673 and ISG15 protein could be a valuable prognostic indicator for pancreatic cancer. The therapies targeting LINC00673 and ISG15 protein signaling pathways are expected to be a potential option for immunotherapy of pancreatic cancer.

The inhibitory effect of curcumin on B16F10 cells and its mechanism / 中国药理学通报

Aim To investigate the effect of curcumin extract on melanin production and melanosome transport, and to explore the possible mechanism of the curcumin extract on microenvironment. Methods (1) B16F10 and HaCaT cells were cultured with different concentrations of curcumin. The proliferation ability was detected by MTT method. Melanin synthesis and tyrosinase activity in B16F10 cells were detected by NaOH pyrolysis method and Oxidation dopamine response in vitro. The expression levels of key proteins were detected by Western blot. (2) B16F10 cells were cultured with different concentrations of ISG15 protein. NaOH pyrolysis method and Oxidation dopamine response in vitro were used to detect melanin synthesis and tyrosinase activity in B16F10 cells. Results Curcumin could directly inhibited tyrosinase activity and melanin production, and inhibit melanocyte migration within a certain concentration range. ISG15 protein could enhance the melanin production, tyrosinase activity. Curcumin could reduce the expression of the ISG15 in HaCaT cell, change the microenvironment of melanocyte, and indirectly inhibit melanin synthesis through ISG15. Conclusions In addition to directly inhibiting melanin synthesis, curcumin can also play an indirect role in inhibiting melanin synthesis by inhibiting the expression of ISG15 protein and altering the microenvironment of melanocytes.

Baricitinib inhibits type Ⅰ IFN-signaling during SARS-CoV-2 infection in vitro / 中国热带医学

@#Abstract: Objective　To explore the antiviral effect of baricitinib in the SARS-CoV-2 infection and influence on cytokine levels. Methods　Calu-3 cells were infected with SARS-CoV-2 at MOI of 0.1, and the levels of inflammatory cytokines (IL-6, IL-8, TNF-α and IL-1β), interferon β (IFN-β) and interferon-stimulated gene, IFIT2 in the infected cells were analyzed by qRT-PCR methods. At the same time, Calu-3 cells were infected with SARS-CoV-2 (MOI=0.1) after being treated with baricitinib for 2 hours. Cells were collected at 0, 24, 36, and 48 hours, and analyzed for the mRNA of the above genes in the drug-treated and untreated groups. Results　The mRNA levels of IL-6, TNF-a, IL-1β, IFN-β and IFIT2 in Calu-3 infected by SARS-CoV-2 cells were increased significantly. These cytokines were increased by nearly 100-fold post-infection 48 h compared with the control (P<0.000 1), and continued to increase with the infection time (P<0.001 or P<0.000 1). The increase of IL-8 mRNA level was not as significant as IL-6, TNF-α, IL-8, IL-1β, but it also showed a 2-4 folds increase. Baricitinib does not affect the level of viral RNA in Calu-3 cells after SARS-CoV-2 infection (P>0.05). However, baricitinib can significantly inhibit the up-regulation of IL-6 and TNF-α levels induced by SARS-CoV-2 infection (5.25-fold and 3.90-fold down-regulation, respectively, P<0.01), and has little effect on the levels of IL-8 and IL-1β . In addition, the drug could also significantly down-regulate the increase in IFN-β and IFIT2 levels caused by viral infection (10.51-fold and 90.78-fold down-regulation, respectively, P<0.000 1). Conclusions　Baricitinib inhibits the release of inflammatory cytokines to some extent, but it drastically down-regulates the expression of interferons and interferon-stimulated genes (ISGs), and has limited antiviral effect on SARS-CoV-2. Considering that interferon signal pathways play important roles on viral infection, caution should be exercised when using baricitinib to treat COVID-19 patients.

Evaluation of interferon-stimulated gene 15 (ISG15) expression in blood neutrophils in beef cattle poisoned by Senecio spp

ABSTRACT: This study aimed to assess liver damage and interferon-stimulated gene 15 (ISG15) blood expression as a consequence of embryonic signaling on maternal recognition of pregnancy in beef cattle presenting natural ingestion of Senecio spp. Epidemiological aspects, as the presence of the plant, associated to gamma glutamyl transferase (GGT) activity can be used as Senecio spp. poisoning diagnosis. Maternal recognition of pregnancy period occurs when the embryo secretes interferon tau (IFNT) to signal its presence to the mother and eventually extend corpus luteum (CL) lifespan. In our study, liver damage was determined by concentration serum GGT, cytological and histopathological examinations. Reproductive status was evaluated by concentration of progesterone, CL diameter and ISG15 mRNA expression on Day 19 following fixed-time artificial insemination (FTAI). Cows were categorized into two groups based on concentration of GGT: Group 1 (GGT 30U/L) and 2 (GGT>31U/L). No difference on body condition scores was observed. All the cows presented liver damage based on cytology and histopathological exams. Cows from the Group 1 had higher pregnancy rate, presenting larger CL diameter and greater concentration of progesterone. Interestingly, ISG15 mRNA expression had no difference between Groups 1 and 2, even presenting difference in pregnancy status. These findings suggest embryonic loss beyond Day 19. It suggests late embryonic mortality may be associated to liver insufficiency. In conclusion, liver injury and/or concentration of GGT does not alter ISG15 expression on blood neutrophils, however cows presenting lower concentration of GGT ( 30U/L) had increased pregnancy status. Therefore, the concentration of GGT allow us to screen liver status and foresee a successful pregnancy in beef cattle.

RESUMO: O objetivo deste estudo foi avaliar a lesão hepática e a expressão sanguínea do gene estimulado por interferon 15 (ISG15) durante a sinalização embrionária, no reconhecimento materno da gestação, em bovinos de corte apresentando ingestão natural de Senecio spp. Fatores epidemiológicos, como a presença da planta, associados à atividade da gama glutamil transferase (GGT) podem ser utilizados como diagnóstico da intoxicação por Senecio spp. O reconhecimento materno da gestação ocorre quando o embrião secreta interferon tau (IFNT) para sinalizar sua presença à mãe. Em nosso estudo, a lesão hepática foi determinada pela concentração sérica de GGT, pelos exames citológicos e histopatológicos. O estado reprodutivo foi avaliado pela concentração de progesterona, diâmetro de corpo lúteo (CL) e expressão de mRNA ISG15 no Dia 19 após a inseminação artificial em tempo fixo (IATF). As vacas foram separadas em dois grupos com base na concentração de GGT sanguíneo: Grupo 1 (GGT 30U/L) e Grupo 2 (GGT>31U/L). Não foi observada nenhuma diferença no escore de condição corporal entre os grupos. Na citologia e nos exames histopatológicos todas as vacas apresentaram lesão hepática. As vacas do Grupo 1 apresentaram maior taxa de prenhez, maior diâmetro do CL e maior concentração de progesterona. Diferente do esperado, a expressão do mRNA ISG15 não foi diferente entre os Grupos 1 e 2, mesmo apresentando diferença na taxa de prenhez. Esses achados sugerem perda embrionária após o Ddia 19. Isso demonstra que a mortalidade embrionária tardia pode estar associada à insuficiência hepática. Dessa forma, conclui-se que a lesão hepática e/ou concentração de GGT não altera a expressão de ISG15 nos neutrófilos sanguíneos, porém vacas com menor concentração de GGT ( 30U/L) apresentaram maiores taxas de prenhez. Assim, a concentração de GGT nos permite avaliar a saúde hepática e prever uma gestação bem-sucedida em bovinos de corte.

Evaluation of interferon-stimulated gene 15 (ISG15) expression in blood neutrophils in beef cattle poisoned by Senecio spp / Avaliação da expressão do gene estimulado por interferon 15 (ISG15) em neutrófilos sanguíneos em bovinos de corte intoxicados por Senecio spp

This study aimed to assess liver damage and interferon-stimulated gene 15 (ISG15) blood expression as a consequence of embryonic signaling on maternal recognition of pregnancy in beef cattle presenting natural ingestion of Senecio spp. Epidemiological aspects, as the presence of the plant, associated to gamma glutamyl transferase (GGT) activity can be used as Senecio spp. poisoning diagnosis. Maternal recognition of pregnancy period occurs when the embryo secretes interferon tau (IFNT) to signal its presence to the mother and eventually extend corpus luteum (CL) lifespan. In our study, liver damage was determined by concentration serum GGT, cytological and histopathological examinations. Reproductive status was evaluated by concentration of progesterone, CL diameter and ISG15 mRNA expression on Day 19 following fixed-time artificial insemination (FTAI). Cows were categorized into two groups based on concentration of GGT: Group 1 (GGT<30U/L) and 2 (GGT>31U/L). No difference on body condition scores was observed. All the cows presented liver damage based on cytology and histopathological exams. Cows from the Group 1 had higher pregnancy rate, presenting larger CL diameter and greater concentration of progesterone. Interestingly, ISG15 mRNA expression had no difference between Groups 1 and 2, even presenting difference in pregnancy status. These findings suggest embryonic loss beyond Day 19. It suggests late embryonic mortality may be associated to liver insufficiency. In conclusion, liver injury and/or concentration of GGT does not alter ISG15 expression on blood neutrophils, however cows presenting lower concentration of GGT (<30U/L) had increased pregnancy status. Therefore, the concentration of GGT allow us to screen liver status and foresee a successful pregnancy in beef cattle.(AU)

O objetivo deste estudo foi avaliar a lesão hepática e a expressão sanguínea do gene estimulado por interferon 15 (ISG15) durante a sinalização embrionária, no reconhecimento materno da gestação, em bovinos de corte apresentando ingestão natural de Senecio spp. Fatores epidemiológicos, como a presença da planta, associados à atividade da gama glutamil transferase (GGT) podem ser utilizados como diagnóstico da intoxicação por Senecio spp. O reconhecimento materno da gestação ocorre quando o embrião secreta interferon tau (IFNT) para sinalizar sua presença à mãe. Em nosso estudo, a lesão hepática foi determinada pela concentração sérica de GGT, pelos exames citológicos e histopatológicos. O estado reprodutivo foi avaliado pela concentração de progesterona, diâmetro de corpo lúteo (CL) e expressão de mRNA ISG15 no Dia 19 após a inseminação artificial em tempo fixo (IATF). As vacas foram separadas em dois grupos com base na concentração de GGT sanguíneo: Grupo 1 (GGT<30U/L) e Grupo 2 (GGT>31U/L). Não foi observada nenhuma diferença no escore de condição corporal entre os grupos. Na citologia e nos exames histopatológicos todas as vacas apresentaram lesão hepática. As vacas do Grupo 1 apresentaram maior taxa de prenhez, maior diâmetro do CL e maior concentração de progesterona. Diferente do esperado, a expressão do mRNA ISG15 não foi diferente entre os Grupos 1 e 2, mesmo apresentando diferença na taxa de prenhez. Esses achados sugerem perda embrionária após o Ddia 19. Isso demonstra que a mortalidade embrionária tardia pode estar associada à insuficiência hepática. Dessa forma, conclui-se que a lesão hepática e/ou concentração de GGT não altera a expressão de ISG15 nos neutrófilos sanguíneos, porém vacas com menor concentração de GGT (<30U/L) apresentaram maiores taxas de prenhez. Assim, a concentração de GGT nos permite avaliar a saúde hepática e prever uma gestação bem-sucedida em bovinos de corte.(AU)

Viperin Differentially Induces Interferon-Stimulated Genes in Distinct Cell Types

Viperin is an IFN-stimulated gene (ISG)-encoded protein that was identified in human primary macrophages treated with IFN-γ and in human primary fibroblasts infected with cytomegalovirus (CMV). This protein plays multiple roles in various cell types. It inhibits viral replication, mediates signaling pathways, and regulates cellular metabolism. Recent studies have shown that viperin inhibits IFN expression in macrophages, while it enhances TLR7 and TLR9-mediated IFN production in plasmacytoid dendritic cells, suggesting that viperin can play different roles in activation of the same pathway in different cell types. Viperin also controls induction of ISGs in macrophages. However, the effect of viperin on induction of ISGs in cell types other than macrophages is unknown. Here, we show that viperin differentially induces ISGs in 2 distinct cell types, macrophages and fibroblasts isolated from wild type and viperin knockout mice. Unlike in bone marrow-derived macrophages (BMDMs), viperin downregulates the expression levels of ISGs such as bone marrow stromal cell antigen-2, Isg15, Isg54, myxovirus resistance dynamin like GTPase 2, and guanylate binding protein 2 in murine embryonic fibroblasts (MEFs) treated with type I or II IFN. However, viperin upregulates expression of these ISGs in both BMDMs and MEFs stimulated with polyinosinic-polycytidylic acid or CpG DNA and infected with murine CMV. The efficiency of viral entry is inversely proportional to the expression levels of ISGs in both cell types. The data indicate that viperin differentially regulates induction of ISGs in a cell type-dependent manner, which might provide different innate immune responses in distinct cell types against infections.

Regulation of Cellular Antiviral Signaling by Modifications of Ubiquitin and Ubiquitin-like Molecules

The initiation of cellular antiviral signaling depends on host pattern-recognition receptors (PRRs)-mediated recognition of viral nucleic acids that are known as classical pathogen-associated molecular patterns (PAMPs). PRRs recruit adaptor proteins and kinases to activate transcription factors and epigenetic modifiers to regulate transcription of hundreds of genes, the products of which collaborate to elicit antiviral responses. In addition, PRRs-triggered signaling induces activation of various inflammasomes which leads to the release of IL-1β and inflammation. Recent studies have demonstrated that PRRs-triggered signaling is critically regulated by ubiquitin and ubiquitin-like molecules. In this review, we first summarize an updated understanding of cellular antiviral signaling and virus-induced activation of inflammasome and then focus on the regulation of key components by ubiquitin and ubiquitin-like molecules.

Caracterización de la condición periodontal de pacientes adultos con obesidad / Characterization of the periodontal condition in obese adults

La periodontitis es una enfermedad inflamatoria que puede evolucionar hasta la pérdida dentaria. En la presente investigación se caracterizó la condición periodontal de 30 adultos con obesidad, atendidos en el Instituto de Investigaciones en Nutrición (INVESNUT) de la Facultad de Ciencias de la Salud, Universidad de Carabobo y de un grupo control conformado por 15 adultos normo-peso. Los adultos fueron evaluados clínica y odontológicamente, mediante antropometría (Talla, Peso, IMC, circunferencia abdominal) y los índices: índice de higiene oral simplificado (IHOS), índice de gingivitis (IG), índice de sangramiento gingival (IGB "gingival bleeding index"). El grupo de estudio mostró diferencias significativas para IG, IGB IHOS con respecto al control, el análisis de correlación de Pearson para las variables IMC, circunferencia abdominal y las variables sobre la condición periodontal (IG, IGB) mostraron asociación positiva y significativa, aunque se no se observó correlación para las variables IMC y circunferencia abdominal con IHOS. Se concluye que los pacientes obesos presentan una peor condición periodontal y que su higiene oral no corresponde con el estado inflamatorio del periodonto...

Periodontitis is an inflammatory disease that may progress to tooth loss. The objective of this research was to characterize the periodontal condition of 30 obese adults treated at the Nutrition Research Institute (INVESNUT), Faculty of Health Sciences, University of Carabobo and of a control group of 15 normal-weight adults. All participants were assessed clinically and dentally by anthropometry (Height, weight, BMI, abdominal circumference) and indexes: simplified oral hygiene index (OHI-S), gingivitis index (GI) gingival bleeding index (GBI). The study group showed significant differences for GI, GBI, OHI-S relative to the control, and a significant positive association (Pearson correlation analysis) for BMI, abdominal circumference, GI and GBI, showed, although no correlation was observed to the variables BMI, abdominal circumference with OHI-S. It is concluded that obese patients have a worse periodontal condition, but oral hygiene does not correspond to the inflammatory state of the periodontium...

Ubiquitin-like proteins and hepatocellular carcinoma / 中国肿瘤临床

Ubiquitin-like proteins are structurally similar to ubiquitin. These proteins are processed, activated, conjugated, and re-leased from conjugates by enzymatic steps that are similar to the corresponding mechanisms for ubiquitin. Ubiquitin-like proteins regu-late a wide array of cellular processes through modification processes, such as nuclear-cytosolic transport, transcriptional regulation, protein stability, response to stress, and progression through the cell cycle. A large number of recent studies have found dysfunctional ubiquitin-like proteins in hepatocellular carcinoma. These proteins are important in tumorigenesis, cell proliferation, apoptosis, and an-giogenesis. Anticancer drug studies revealed that regulating protein modification by using ubiquitin-like proteins may alter the anti-tu-mor effects of chemotherapy and thus influence the chemosensitivity of hepatocellular carcinoma. Results indicate that ubiquitin-like proteins may become a new target for cancer therapy. The mechanism of ubiquitin-like proteins in tumorigenesis and hepatocellular car-cinoma progression is of great significance in the diagnosis and treatment of hepatocellular carcinoma.

ISG15: leading a double life as a secreted molecule

ISG15 is a well-known intracellular ubiquitin-like molecule involved in ISGylation. However, a recent study has revived the notion first put forward two decades ago that ISG15 is also a secreted molecule. Human neutrophils, monocytes and lymphocytes can release ISG15, even though this protein has no detectable signal peptide sequence. ISG15 has also been found in the secretory granules of granulocytes. The mechanism underlying ISG15 secretion is unknown. Secreted ISG15 acts on at least T and natural killer (NK) lymphocytes, in which it induces interferon (IFN)-gamma production. However, the mechanism by which ISG15 stimulates these cells also remains unclear. ISG15 and IFN-gamma seem to define an innate circuit that operates preferentially, but not exclusively, between granulocytes and NK cells. Inherited ISG15 deficiency is associated with severe mycobacterial disease in both mice and humans. This infectious phenotype probably results from the lack of secreted ISG15, because patients and mice with other inborn errors of IFN-gamma immunity also display mycobacterial diseases. In addition to raising mechanistic issues, the studies described here pave the way for clinical studies of various aspects, ranging from the use of recombinant ISG15 in patients with infectious diseases to the use of ISG15-blocking agents in patients with inflammatory diseases.

Função do interferon-tau durante o reconhecimento materno da gestação em ruminantes / The role of interferon-tau during maternal recognition of pregnancy in ruminants

O reconhecimento materno da gestação é o período em que o concepto sinaliza sua presença para a mãe. Em ruminantes, este período coincide com o alongamento do embrião e a máxima produção de interferon-tau (IFNT). O IFNT produzido pelo concepto age via parácrina no útero inibindo a expressão dos receptores de estrógenos (ESR1) e de ocitocina (OXTR) no epitélio luminal do endométrio, evitando, assim, a liberação de pulsos luteolíticos de prostaglandina F2 alfa (PGF2 ), hormonio responsável pelo início da luteólise. Além da sua ação durante o reconhecimento materno da gestação em ruminantes, o IFNT aumenta a expressão de vários genes estimulados por interferons (ISGs) no útero, no corpo lúteo (CL) e em células sanguíneas. Estudos recentes demonstraram que o IFNT possui ação endócrina no CL ovino e também estende o ciclo estral (pseudo gestação) além do dia 32 após a infusão de IFNT recombinante ovino (roIFNT) na veia uterina. A comprovação da saída de IFNT do útero pela veia uterina sugere que a ação endócrina do IFNT possa ser um mecanismo complementar ao mecanismo intrauterino de reconhecimento materno da gestação. A ação direta do IFNT em tecidos extrauterinos estimula a expressão de ISGs que, no CL, podem estar envolvidos com a resistência luteal à ação luteolítica da PGF2a.

Maternal recognition of pregnancy is the period when the conceptus signals its presence to the dam. In ruminants, it requires conceptus elongation, which coincides with maximum production of interferon-tau (IFNT). Conceptus IFNT acts in a paracrine manner silencing estrogen receptor alpha (ESR1) and oxytocin receptor (OXTR) in the luminal epithelium, thus preventing luteolytic prostaglandin F2 alpha (PGF2 ) pulses. Besides its role during maternal recognition of pregnancy, IFNT induces the expression of several interferon stimulated genes (ISGs) in the endometrium, corpus luteum (CL) and blood cells. Recently, it was suggested an endocrine role for IFNT during the period of maternal recognition of pregnancy in sheep. It was demonstrated that infusion of IFNT into the uterine vein can extend the estrous cycle beyond 32 days. This direct action of IFNT in extrauterine tissues induces ISGs expression, which might be involved in the rescue of the CL from the luteolytic effects of PGF2 pulses.

Cloning, Eukaryotic Expression of Human ISG20 and Preliminary Study on the Effect of Its Anti-HBV / 华中科技大学学报（医学）（英德文版）

Human ISG20 gene was cloned and the effect of its anti-HBV was primarily studied. The ISG20 gene was amplified from HeLa cells by RT-PCR and recombinant vector expressing ISG20 was constructed by genetic engineering. The overexpression of ISG20 in HepG2 cells was detected by Western blot and the levels of secretion of HBs antigen and HBe antigen tested by ELISA. The results showed that: (1) Sequence of ISG20 cloned was consistent to that published in Genebank; (2) Recombinant vector expressing ISG20 could be expressed in HepG2 cells by transfection; (3) The overexpression of ISG20 protein could reduce the levels of the secretion of HBs antigen and HBe an-tigen in transfected HepG2 cells. It was suggested that the overexpression of recombinant ISG20 in culture cells could reduce the synthesis of HBV proteins.

Ubiquitin-like Protein ISG15 and Its Role in Innate Immunity / 生物化学与生物物理进展

Virus infection or interferon can stimulate robust expression of the protein ISG15 that is encoded by interferon stimulated gene 15, which was the first unbiquitin-like molecule identified two decades ago. While ubiquitin and its many important functions have been well established, the functions of ISG15 and its post-translational conjugation are still largely unknown . Recently, some specific enzymes have been identified to be involved in the ISG15 modification system, suggests that ISG15 and its modification system play important roles in the innate immune response and regulation of interferon signaling. The history of ISG15 discovery and its biochemical characterization were briefly introduced. Then such topics as the ISG15 gene expression and the ISG15 modification will be focued on, and finally summarize new findings which have implications for ISG15 and its modification system in immunology and interferon signal transduction were summarized.

Expression of interferon-stimulated gene ISG20 and investigation of its antiviral effect on HCV replicon / 中国免疫学杂志

Objective:To study the antiviral effect of ISG20 on HCV replicon.Methods:Wild type ISG20/mutated ISG20 cDNAs were obtained by RT-PCR/two step-PCR directed mutagenesis, and wild type ISG20 and dominant negative mutated ISG20 mammal expression vectors were consuructed. The constructed pISG20wt and pISG20m expressing vectors were transfected into Huh7 cells or Huh7 cells containing HCV replicon to investigate its effects on HCV replicon replication.Results:The ISG20wt/ISG20m expression vectors were constructed and the expressions of these two vectors were confirmed at both mRNA and protein levels. The effects of ISG20wt on HCV replicon replication were evaluated by Northern blot and Western blot. The results showed that expression of ISG20wt had significant inhibitory effect on HCV RNA replication.Conclusion:ISG20 participates in the anti-HCV action of IFN-? on HCV replicon system.