RESUMEN
Upon infection, HIV disseminates throughout the human body within 1-2 weeks. However, its early cellular targets remain poorly characterized. We used a single-cell approach to retrieve the phenotype and TCR sequence of infected cells in blood and lymphoid tissue from individuals at the earliest stages of HIV infection. HIV initially targeted a few proliferating memory CD4+ T cells displaying high surface expression of CCR5. The phenotype of productively infected cells differed by Fiebig stage and between blood and lymph nodes. The TCR repertoire of productively infected cells was heavily biased, with preferential infection of previously expanded and disseminated clones, but composed almost exclusively of unique clonotypes, indicating that they were the product of independent infection events. Latent genetically intact proviruses were already archived early in infection. Hence, productive infection is initially established in a pool of phenotypically and clonotypically distinct T cells, and latently infected cells are generated simultaneously.
Asunto(s)
Infecciones por VIH , VIH-1 , Infección Latente , Humanos , Linfocitos T CD4-Positivos/metabolismo , VIH-1/genética , Infección Latente/metabolismo , Infección Latente/patología , Receptores de Antígenos de Linfocitos T/metabolismo , Latencia del VirusRESUMEN
OBJECTIVE: To evaluate the quantity of tumor-associated macrophages (TAMs) in cases of Hodgkin Lymphoma of classical type (cHL), and to reveal possible associations between TAM intensity and latent Epstein-Barr virus (EBV) infection, overall survival, progression-free survival, prognostic indices, and clinicopathological parameters. MATERIALS AND METHODS: A total 46 cases of cHL with complete clinical records were selected and re-evaluated histopathologically. Staining for CD68 (PG-M1; KP1 clones) and CD163 was evaluated and the cut-off values were defined. Also, all cases were evaluated using the chromogen in situ hybridization (CISH) method with EBER (Epstein-Barr virus-encoded RNA) probes for the presence of possible EBV infection. RESULTS: It was found that high expression levels of PG-M1 and high International Prognostic Scores (IPS) were associated with shortened overall survival (p=0.047, p=0.013). Cases with 2 or less areas of nodal region involvement were observed to have longer progression-free survival period (p=0.043). Higher expression levels of CD68 PG-M1, CD68 KP1, and CD163 were found to show significant associations with the presence of some clinical parameters such as the presence of B symptoms, spleen involvement, and the presence of EBV infection. CONCLUSIONS: Our findings suggest that increase of PG-M1+ TAM is associated with shortened overall survival, while higher expressions of all immunohistochemical markers are statistically significantly associated with the presence of EBV infection and clinical parameters mentioned above. These findings indicate that highlighting the TAM rate via macrophage markers in cases of cHL could be helpful in determining the prognostic risk groups and the relevant results should be mentioned in pathology reports.
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Antígenos CD/análisis , Antígenos de Diferenciación Mielomonocítica/análisis , Biomarcadores de Tumor/análisis , Infecciones por Virus de Epstein-Barr/inmunología , Herpesvirus Humano 4/genética , Enfermedad de Hodgkin/inmunología , Infección Latente/inmunología , Receptores de Superficie Celular/análisis , Macrófagos Asociados a Tumores/inmunología , Infecciones por Virus de Epstein-Barr/patología , Infecciones por Virus de Epstein-Barr/terapia , Infecciones por Virus de Epstein-Barr/virología , Femenino , Enfermedad de Hodgkin/patología , Enfermedad de Hodgkin/terapia , Enfermedad de Hodgkin/virología , Humanos , Inmunohistoquímica , Hibridación in Situ , Infección Latente/patología , Infección Latente/terapia , Infección Latente/virología , Masculino , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Supervivencia sin Progresión , Factores de Riesgo , Factores de Tiempo , Macrófagos Asociados a Tumores/patología , Macrófagos Asociados a Tumores/virologíaRESUMEN
BACKGROUND: Genetics have provided hints on potential molecular pathways involved in neurodegenerative diseases (NDD). However, the number of cases caused exclusively by genetic alterations is low, suggesting an important contribution of environmental factors to NDDs. Among these factors, viruses like herpes simplex viruses (HSV-2), capable of establishing lifelong infections within the nervous system (NS), are being proposed to have a role in NDDs. Despite promising data, there is a significant lack of knowledge on this and an urgent need for more research. METHODS: We have set up a mouse model to study HSV latency and its associated neuroinflammation in the spinal cord. The goal of this model was to observe neuroinflammatory changes caused by HSV latent infections, and if those changes were similar to alterations observed in the spinal cord of amyotrophic lateral sclerosis (ALS) patients. RESULTS: In infected spinal cords, we have observed a strong leukocyte infiltration and a severe alteration of microglia close to motor neurons. We have also analyzed ALS-related proteins: we have not found changes in TDP-43 and Fus in neurons, but interestingly, we have found decreased protein levels of C9orf72, of which coding gene is severely altered in some familial forms of ALS and is critical for microglia homeostasis. CONCLUSIONS: Latent infection of HSV in the spinal cord showed altered microglia and leukocyte infiltration. These inflammatory features resembled to those observed in the spinal cord of ALS patients. No changes mimicking ALS neuropathology, such as TDP-43 cytoplasmic inclusions, were found in infected spinal cords, but a decrease in protein levels of C9orf72 was observed. Then, further studies should be required to determine whether HSV-2 has a role in ALS.
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Esclerosis Amiotrófica Lateral/metabolismo , Proteína C9orf72/metabolismo , Herpes Genital/metabolismo , Herpesvirus Humano 2/aislamiento & purificación , Infección Latente/metabolismo , Médula Espinal/metabolismo , Esclerosis Amiotrófica Lateral/inmunología , Animales , Proteína C9orf72/análisis , Modelos Animales de Enfermedad , Femenino , Herpes Genital/inmunología , Herpes Genital/patología , Herpesvirus Humano 2/inmunología , Infección Latente/inmunología , Infección Latente/patología , Ratones , Ratones Endogámicos BALB C , Médula Espinal/patología , Médula Espinal/virología , Vagina/inmunología , Vagina/metabolismo , Vagina/patología , Vagina/virologíaRESUMEN
Generating oxidative stress is a critical mechanism by which host cells defend against infection by pathogenic microorganisms. Radiation resistance is a critical problem in radiotherapy against cancer. Epstein-Barr virus (EBV) is a cancer-causing virus and its reactivation plays an important role in the development of EBV-related tumors. This study aimed to explore the inner relationship and regulatory mechanism among oxidative stress, EBV reactivation, and radioresistance and to identify new molecular subtyping models and treatment strategies to improve the therapeutic effects of radiotherapy. Methods: ROS, NADP+/NADPH, and GSSG/GSH were detected to evaluate the oxidative stress of cells. 8-OHdG is a reliable oxidative stress marker to evaluate the oxidative stress in patients. Its concentration in serum was detected using an ELISA method and in biopsies was detected using IHC. qPCR array was performed to evaluate the expression of essential oxidative stress genes. qPCR, Western blot, and IHC were used to measure the level of EBV reactivation in vitro and in vivo. A Rta-IgG ELISA kit and EBV DNA detection kit were used to analyze the reactivation of EBV in serum from NPC patients. NPC tumor tissue microarrays was used to investigate the prognostic role of oxidative stress and EBV reactivation. Radiation resistance was evaluated by a colony formation assay. Xenografts were treated with NAC, radiation, or a combination of NAC and radiation. EBV DNA load of tumor tissue was evaluated using an EBV DNA detection kit. Oxidative stress, EBV reactivation, and the apoptosis rate in tumor tissues were detected by using 8-OHdG, EAD, and TUNEL assays, respectively. Results: We found that EBV can induce high oxidative stress, which promotes its reactivation and thus leads to radioresistance. Basically, EBV caused NPC cells to undergo a process of 'Redox Resetting' to acquire a new redox status with higher levels of ROS accumulation and stronger antioxidant systems by increasing the expression of the ROS-producing enzyme, NOX2, and the cellular master antioxidant regulator, Nrf2. Also, EBV encoded driving protein LMP1 promotes EBV reactivation through production of ROS. Furthermore, high oxidative stress and EBV reactivation were positively associated with poor overall survival of patients following radiation therapy and were significant related to NPC patients' recurrence and clinical stage. By decreasing oxidative stress using an FDA approved antioxidant drug, NAC, sensitivity of tumors to radiation was increased. Additionally, 8-OHdG and EBV DNA could be dual prognostic markers for NPC patients. Conclusions: Oxidative stress mediates EBV reactivation and leads to radioresistance. Targeting oxidative stress can provide therapeutic benefits to cancer patients with radiation resistance. Clinically, we, for the first time, generated a molecular subtyping model for NPC relying on 8-OHdG and EBV DNA level. These dual markers could identify patients who are at a high risk of poor outcomes but who might benefit from the sequential therapy of reactive oxygen blockade followed by radiation therapy, which provides novel perspectives for the precise treatment of NPC.
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Quimioradioterapia/métodos , Infecciones por Virus de Epstein-Barr/terapia , Depuradores de Radicales Libres/administración & dosificación , Carcinoma Nasofaríngeo/terapia , Neoplasias Nasofaríngeas/terapia , Acetilcisteína/administración & dosificación , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/sangre , Biopsia , Línea Celular Tumoral , ADN Viral/sangre , Infecciones por Virus de Epstein-Barr/sangre , Infecciones por Virus de Epstein-Barr/mortalidad , Infecciones por Virus de Epstein-Barr/virología , Femenino , Estudios de Seguimiento , Disulfuro de Glutatión/sangre , Disulfuro de Glutatión/metabolismo , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/aislamiento & purificación , Herpesvirus Humano 4/patogenicidad , Humanos , Infección Latente/sangre , Infección Latente/patología , Infección Latente/terapia , Infección Latente/virología , Masculino , Persona de Mediana Edad , Carcinoma Nasofaríngeo/sangre , Carcinoma Nasofaríngeo/mortalidad , Carcinoma Nasofaríngeo/virología , Neoplasias Nasofaríngeas/sangre , Neoplasias Nasofaríngeas/mortalidad , Neoplasias Nasofaríngeas/virología , Nasofaringe/patología , Nasofaringe/virología , Estrés Oxidativo/efectos de los fármacos , Selección de Paciente , Pronóstico , Supervivencia sin Progresión , Tolerancia a Radiación/efectos de los fármacos , Especies Reactivas de Oxígeno , Carga Viral , Proteínas de la Matriz Viral/metabolismo , Activación Viral/efectos de los fármacos , Ensayos Antitumor por Modelo de Xenoinjerto , Adulto JovenRESUMEN
Rationale: Epstein-Barr virus (EBV) is found in ~7% of gastric carcinoma cases worldwide, and all tumour cells harbour the clonal EBV genome. EBV can regulate pathways and protein expression to induce gastric carcinoma; however, the molecular mechanism underlying EBV-associated gastric carcinoma (EBVaGC) remains elusive. Methods: GEO microarray and molecular experiments were performed to compare CXCR4 expression between EBV-positive and EBV-negative gastric carcinoma (EBVnGC). Transfections with LMP2A plasmid or siRNA were carried out to assess the role of LMP2A in CXCR4 expression. The effects and mechanisms of CXCR4 on cell autophagy were analysed in vitro using molecular biological and cellular approaches. Additionally, we also determined the regulatory role of CXCR4 in latent EBV infection. Results: CXCR4 expression was significantly upregulated in EBVaGC tissues and cell lines. LMP2A could induce AKT phosphorylation to increase NRF1 expression, thereby binding to the CXCR4 promoter to increase its transcriptional level. Moreover, CXCR4 promoted ZEB1 expression to upregulate ATG7 synthesis, which could then activate autophagy. Moreover, CXCR4 increased the number of cells entering the G2/M phase and inhibited cell apoptosis via the autophagy pathway. Finally, CXCR4 knockdown was associated with elevated BZLF1 expression, but this effect was not influenced by autophagy. Conclusions: Our data suggested new roles for CXCR4 in autophagy and EBV replication in EBVaGC, which further promoted cell survival and persistent latent infection. These new findings can lead to further CXCR4-based anticancer therapy.