HCV spontaneous clearers showed low senescence profile in people living with HIV under long ART.

Coinfection with hepatitis C virus (HCV) and human immunodeficiency virus (HIV) increases immune activation, inflammation, and oxidative stress that could lead to premature senescence. Different HCV infections, either acute or chronic infection, could lead to distinct premature cellular senescence in people living with HIV (PLWHIV). Observational study in 116 PLWHIV under antiretroviral treatment with different HCV status: (i) n = 45 chronically infected with HCV (CHC); (ii) n = 36 individuals who spontaneously clarify HCV (SC); (iii) n = 35 HIV controls. Oxidative stress biomarkers were analyzed at lipid, DNA, protein, and nitrates levels, as well as antioxidant capacity and glutathione reductase enzyme. Replicative senescence was evaluated by relative telomere length (RTL) measurement. Additionally, 26 markers of Senescence-Associated Secretory Phenotype (SASP) were analyzed by multiplex immunoassays (Luminex xMAP technology). Differences were evaluated by generalized linear model (GLMs) adjusted by most significant covariates. The SC group had a senescence signature similar to the HIV control group and slightly lower SASP levels. However, significant differences were observed with respect to the CHC group, where an increase in the nitrate concentration [adjusted arithmetic mean ratio, aAMR = 1.73 (1.27-2.35), p < 0.001, q = 0.009] and the secretion of 13 SASP-associated factors [granulocyte macrophage colony-stimulating factor (GM-CSF), interferon-ß, interleukin (IL)-1ß, IL-2, IL-8, IL-13, tumor necrosis factor (TNF)-α, IL-1α, IL-1RA, IL-7, IL-15, C-X-C motif chemokine ligand 10 (IP-10), stem cell factor (SCF); q < 0.1)] was detected. The CHC group also showed higher values of IL-1α, IP-10, and placental growth factor 1 (PIGF-1) than HIV controls. The SC group showed a slightly lower senescence profile than the HIV group, which could indicate a more efficient control of viral-induced senescence due to their immune strengths. Chronic HCV infection in PLWHIV led to an increase in nitrate and elevated SASP biomarkers favoring the establishment of viral persistence.

Dynamics of cellular senescence markers after HCV elimination spontaneously or by DAAs in people living with HIV.

BACKGROUND: We identified that acute or chronic Hepatitis C (HCV) infection in people living with HIV (PLWHIV) results in different senescence profiles. However, variations in these profiles after HCV elimination, spontaneously or with direct-acting antivirals (DAAs), remain unclear. METHODS: Longitudinal observational study (48 weeks) in 70 PLWHIV: 23 PLWHIV with active HCV-chronic infection (CHC) before and after HCV eradication with DAAs, 12 PLWHIV who spontaneously clarify the HCV (SC), and 35 controls (HIV). Oxidative stress was quantified at DNA, lipid, protein, and nitrate levels, as well as the antioxidant capacity and glutathione enzyme. The replicative senescence was evaluated by relative telomere length measurement by PCR and twenty-six factors related to Senescence-Associated Secretory Phenotype (SASP) were characterized by Luminex. Differences in senescence markers was evaluated by generalized linear models. RESULTS: During follow-up, the SC group achieved a significant improvement in glutathione enzyme and lipid peroxidation. The secretion of SASP markers increased but was still lower than that of the HIV group. Overall, the CHC group reduced the levels of oxidative stress and SASP markers to levels like those of the HIV group. No significant differences in telomere shortening were observed between groups. CONCLUSIONS: As the time since spontaneous resolution of HCV infection increased, patients had an improved senescence profile compared to the HIV group. Elimination of chronic HCV infection by DAAs led to a partial improvement of the senescent profile by restoring oxidative stress levels. However, although some SASP markers reached levels like those of the HIV group, others remained altered.

Dynamics of HIV Reservoir and HIV-1 Viral Splicing in HCV-Exposed Individuals after Elimination with DAAs or Spontaneous Clearance.

Background: Although human immunodeficiency virus type 1 (HIV-1) reservoir size is very stable under antiretroviral therapy (ART), individuals exposed to the Hepatitis C virus (HCV) (chronically coinfected and spontaneous clarifiers) show an increase in HIV reservoir size and in spliced viral RNA, which could indicate that the viral protein regulator Tat is being more actively synthesized and, thus, could lead to a higher yield of new HIV. However, it is still unknown whether the effect of HCV elimination with direct-acting antivirals (DAAs) could modify the HIV reservoir and splicing. Methods: This longitudinal study (48 weeks' follow-up after sustained virological response) involves 22 HIV+-monoinfected individuals, 17 HIV+/HCV- spontaneous clarifiers, and 24 HIV+/HCV+ chronically infected subjects who eliminated HCV with DAAs (all of them aviremic, viral load < 50). Viral-spliced RNA transcripts and proviral DNA copies were quantified by qPCR. Paired samples were analyzed using a mixed generalized linear model. Results: A decrease in HIV proviral DNA was observed in HIV+/HCV- subjects, but no significant differences were found for the other study groups. An increased production of multiple spliced transcripts was found in HIV+ and HIV+/HCV+ individuals. Conclusions: We conclude that elimination of HCV by DAAs was unable to revert the consequences derived from chronic HCV infection for the reservoir size and viral splicing, which could indicate an increased risk of rapid HIV-reservoir reactivation. Moreover, spontaneous clarifiers showed a significant decrease in the HIV reservoir, likely due to an enhanced immune response in these individuals.

903 Protein Saver cards: the best alternative for dried blood spot storage at room temperature for HCV RNA.

Hepatitis C virus (HCV) infection remains a global health problem, detected only in the early stages by molecular tests. Molecular tests detect HCV RNA, which is very prone to degradation by ribonucleases, reason why blood samples must be transported and stored at - 20 °C, or even - 70 °C for long-term storage. Flinders Technology Associates (FTA) cards are a useful sampling collecting device for dry blood spot (DBS) storage, especially for low and middle-income countries (LMIC). In this study, we analyzed viral HCV RNA integrity for long-term storage at room temperature compared to - 20 °C using two different types of cards for DBS: FTA Classic and 903 Protein Saver cards. For this purpose, DBS were prepared on these cards using blood or plasma samples from HCV infected patients, and samples were analysed by conventional RT-PCR. Our results showed that 903 Protein Saver cards are the best and cheapest alternative for DBS storage at room temperature. In these conditions, we found that HCV RNA integrity lasted for up to 9 months.

Persistent low-Level viremia in persons living with HIV undertreatment: An unresolved status.

Antiretroviral therapy (ART) allows suppressed viremia to reach less than 50 copies/mL in most treated persons living with HIV (PLWH). However, the existence of PLWH that show events of persistent low-level viremia (pLLV) between 50 and 1000 copies/mL and with different virological consequences have been observed. PLLV has been associated with higher virological failure (VF), viral genotype resistance, adherence difficulties and AIDS events. Moreover, some reports show that pLLV status can lead to residual immune activation and inflammation, with an increased risk of immunovirological failure and a pro-inflammatory cytokine level which can lead to a higher occurrence of non-AIDS defining events (NADEs) and other adverse clinical outcomes. Until now, however, published data have shown controversial results that hinder understanding of the true cause(s) and origin(s) of this phenomenon. Molecular mechanisms related to viral reservoir size and clonal expansion have been suggested as the possible origin of pLLV. This review aims to assess recent findings to provide a global view of the role of pLLV in PLWH and the impact this status may cause on the clinical progression of these patients.

Different HCV Exposure Drives Specific miRNA Profile in PBMCs of HIV Patients.

Micro RNAs (miRNAs) are essential players in HIV and HCV infections, as both viruses modulate cellular miRNAs and interact with the miRNA-mediated host response. We aim to analyze the miRNA profile of HIV patients with different exposure to HCV to explore specific signatures in the miRNA profile of PBMCs for each type of infection. We massively sequenced small RNAs of PBMCs from 117 HIV+ infected patients: 45 HIV+ patients chronically infected with HCV (HIV/HCV+), 36 HIV+ that spontaneously clarified HCV after acute infection (HIV/HCV-) and 36 HIV+ patients without previous HCV infection (HIV). Thirty-two healthy patients were used as healthy controls (HC). Differential expression analysis showed significantly differentially expressed (SDE) miRNAs in HIV/HCV+ (n = 153), HIV/HCV- (n = 169) and HIV (n = 153) patients. We found putative dysregulated pathways, such as infectious-related and PI3K signaling pathways, common in all contrasts. Specifically, putatively targeted genes involved in antifolate resistance (HIV/HV+), cancer-related pathways (HIV/HCV-) and HIF-signaling (HIV) were identified, among others. Our findings revealed that HCV strongly influences the expression profile of PBMCs from HIV patients through the disruption of its miRNome. Thus, different HCV exposure can be identified by specific miRNA signatures in PBMCs.

Hepatitis C Virus Influences HIV-1 Viral Splicing in Coinfected Patients.

Coinfection with hepatitis C virus (HCV) influences HIV reservoir size. However, it is unknown whether this coinfection also induces a higher provirus transcription. Viral transcription is promoted by synergy between cellular factors such as NF-κB and the viral regulator Tat. The impact of HCV coinfection on HIV provirus transcription was analyzed in resting (r)CD4 T+ cells (CD3+CD4+CD25-CD69-HLADR-) and rCD4 T cells-depleted PBMCs (rCD4 T- PBMCs) from a multicenter cross-sectional study of 115 cART-treated HIV patients: 42 HIV+/HCV+ coinfected individuals (HIV+/HCV+), 34 HIV+ patients with HCV spontaneous clearance (HIV+/HCV-) and 39 HIV patients (HIV+). Viral transcription was assessed in total RNA through the quantification of unspliced, single spliced, and multiple spliced viral mRNAs by qPCR. Linear correlations between viral reservoir size and viral splicing were determined. A 3-fold increase of multiple spliced transcripts in rCD4 T+ cells of HIV+/HCV+ patients was found compared to HIV+ individuals (p < 0.05). As Tat is synthesized by multiple splicing, the levels of Tat were also quantified in these patients. Significant differences in single and multiple spliced transcripts were also observed in rCD4 T- PBMCs. Levels of multiple spliced mRNAs were increased in rCD4 T+ cells isolated from HIV+/HCV+ subjects, which could indicate a higher Tat activity in these cells despite their resting state.