Viruses

The concentration of extracellular vesicles (EVs) in the peripheral blood of COVID-19 patients is increased. Nevertheless, their potential role in the transmission of infection remains unclear. This study was performed to determine whether EVs produced by the sub-genomic replicon system developed in Baby Hamster Kidney (BHK-21) cells could transfer SARS-CoV-2 replicon RNA, leading to the establishment of a viral replication system in human cells. Purified EVs from the SARS-CoV-2 sub-genomic replicon cell line BHK-21 were cultured with a naive human cell line. The success of EV-mediated transfer of SARS-CoV-2 replicon RNA and its productive replication was assessed using G-418 selection, a luciferase assay, immunostaining, and Western blot. We found that the A549 cell line cultured with EVs isolated from SARS-CoV-2 BHK-21 replicon cells developed G-418-resistant cell colonies. SARS-COV-2 RNA replication in A549 cells was confirmed by nano luciferase, Nsp1 protein. SARS-CoV-2 RNA replication causes massive morphological changes. Treatment of cells with the FDA-approved Paxlovid demonstrated a dose-dependent inhibition of viral replication. We isolated two human epithelial cell lines (gastrointestinal and neuroblastoma) and one vascular endothelial cell line that stably support high-level replication of SARS-CoV-2 sub-genomic RNA. Viral elimination did not revert the abnormal cellular shape, vesicle accumulation, syncytia formation, or EV release. Our study’s findings highlight the potential implications of EV-mediated transfer of replicon RNA to permissive cells. The replicon model is a valuable tool for studying virus-induced reversible and irreversible cellular reprogramming, as well as for testing novel therapeutic strategies for SARS-CoV-2.

Studying the incidence of Kaposi sarcoma in relation to key variables can guide targeted research and subtype-specific clinical interventions. We reviewed the records of all patients who visited our hospital’s dermatology outpatient clinic, and patients who were clinically and histopathologically diagnosed with Kaposi sarcoma were included in the study. The age, gender, lesion location, anti-HIV test results, and comorbidities of the patients were recorded. Thirty-three patients with Kaposi sarcoma were identified. The male/female ratio was 2.7:1. The Kaposi sarcoma lesions were statistically significantly more prevalent in the lower extremities of HIV-negative patients (p = 0.005). Receiver operating characteristic (ROC) curve analysis identified 59 years as the optimal age cutoff for distinguishing between HIV-positive and HIV-negative patients. Anti-HIV positivity was significantly higher in individuals aged 59 and younger compared to those aged 60 and older (p < 0.001). To the best of our knowledge, this is the first study to demonstrate a statistically significant higher prevalence of lower extremity lesions among HIV-negative patients and to identify 59 years as the optimal age cutoff for distinguishing between HIV-positive and HIV-negative Kaposi sarcoma patients using ROC curve analysis. The age-related patterns observed in this study warrant further investigation.

Oncolytic coxsackievirus B3 (oCVB3) strain PD-H has shown potent oncolytic efficacy and a remarkable safety profile in the treatment of colorectal cancer in vivo after intratumoral (i.t.) injection. In this study, we investigated the safety and efficiency of PD-H following intravenous (i.v.) virus administration. When injected i.v. into Balb/C mice bearing subcutaneous Colon-26 tumors, PD-H led to slightly reduced tumor progression and a significant increase in animal survival, but it also caused multi-organ infection and tissue damage. To improve the safety profile of PD-H, we inserted microRNA target sites (miR-TS) of the heart-specific miR-1, pancreas-specific miR-375, liver-specific miR-122, and brain-specific miR-124 or the tumor-suppressor miR-145 into the genome of PD-H and generated the viruses PD-622TS and PD-145TS. Both viruses replicated similarly and induced cytotoxicity comparable to that of PD-H in the colorectal carcinoma cell lines Colon-26 and CT-26Luc. Their replication was inhibited in HEK293T cells transiently transfected with the cognate microRNAs. In vivo, i.v. administration of PD-145TS and PD-622TS to healthy Balb/C mouse resulted in significantly lower viral titers in the organs of mice and led to significantly less-intense pathological alterations compared to PD-H. PD-622TS injected i.v. into Balb/C mice with CT-26Luc-induced peritoneal carcinomatosis did not induce off-target alterations in normal organs, but it failed to induce a therapeutic effect. These data indicate that PD-H or microRNA-regulated PD derivatives exhibit only limited therapeutic efficacy following i.v. injection in colorectal tumor-bearing mice. However, the newly engineered microRNA-regulated PD-H variants demonstrate improved safety profiles.