When confronted with poor oxygenation, cells adapt by activating survival signaling pathways, including the oxygen-sensitive transcriptional regulators called hypoxia-inducible factor alphas (HIF-αs). We report here that HIF-1α also regulates the life cycle of Epstein-Barr virus (EBV). Incubation of EBV-positive gastric carcinoma AGS-Akata and SNU-719 and Burkitt lymphoma Sal and KemIII cell lines with a prolyl hydroxylase inhibitor, L-mimosine or deferoxamine, or the NEDDylation inhibitor MLN4924 promoted rapid and sustained accumulation of both HIF-1α and lytic EBV antigens. ShRNA knockdown of HIF-1α significantly reduced deferoxamine-mediated lytic reactivation. HIF-1α directly bound the promoter of the EBV primary latent-lytic switch BZLF1 gene, Zp, activating transcription via a consensus hypoxia-response element (HRE) located at nt -83 through -76 relative to the transcription initiation site. HIF-1α did not activate transcription from the other EBV immediate-early gene, BRLF1. Importantly, expression of HIF-1α induced EBV lytic-gene expression in cells harboring wild-type EBV, but not in cells infected with variants containing base-pair substitution mutations within this HRE. Human oral keratinocyte (NOK) and gingival epithelial (hGET) cells induced to differentiate by incubation with either methyl cellulose or growth in organotypic culture accumulated both HIF-1α and Blimp-1α, another cellular factor implicated in lytic reactivation. HIF-1α activity also accumulated along with Blimp-1α during B-cell differentiation into plasma cells. Furthermore, most BZLF1-expressing cells observed in lymphomas induced by EBV in NSG mice with a humanized immune system were located distal to blood vessels in hypoxic regions of the tumors. Thus, we conclude that HIF-1α plays central roles in both EBV’s natural life cycle and EBV-associated tumorigenesis. We propose that drugs that induce HIF-1α protein accumulation are good candidates for development of a lytic-induction therapy for treating some EBV-associated malignancies.

In this large population-based cohort, CNLDO occurred in one in nine live births with no gender predilection. Prematurity and Caucasian race were associated with the development of CNLDO. Mucopurulent discharge was a much more common feature than tearing at initial presentation.

Importance: Although the overall rate of spontaneous resolution in congenital nasolacrimal duct obstruction (CNLDO) and efficacy of probing have been documented in the literature, the optimal timing of intervention has not been established. Objective: To report new findings regarding spontaneous resolution in a large cohort of children with CNLDO Design, Setting, and Participants: The medical records of 1998 consecutive infants diagnosed with CNLDO from January 1, 1995, through December 31, 2004, while residing in Olmsted County, Minnesota were retrospectively reviewed. Data were analyzed between January 1, 2015, and January 2017. Main Outcome(s) and Measure(s): Rate of spontaneous resolution over time and by sex. Results: The cohort, diagnosed at a median age of 1.2 months (interquartile range, 0.4–3.6), was 48% girls and 89% white. Among the 1998 cases, 1669 (83.5%) spontaneously resolved, 289 (14.5%) underwent treatment, and the remaining 40 (2.0%) were lost to follow-up. Of the 1958 followed infants, 47.3% spontaneously resolved by 3 months of age, 66.4% by 6, 75.7% by 9, and 78.4% by 12 months. The rate of resolution was 35% faster (95% CI=23%−47%; p<0.001) at <1 month vs 3 months, 43% faster (95% CI=27%−43%; p<0.001) at 3 months vs. 6 months, 39% faster (95% CI=16%−64%; p<0.001) at 6 months vs. 9 months, and insignificantly different at 9 vs. 12 months (HR=0.99; 95% CI=0.80–1.22; p=0.78). Males resolved 24% faster than females (95% CI=13%−37%; p<0.001), and unilateral obstructions resolved 16% faster than bilateral (95% CI=5%−29%; p=0.005) ones. Children probed at 15months or older had decreased odds of resolution after probing (OR=0.11; 95% CI=0.01–0.89; p=0.04) compared to children probed at 12–14 months of age. Conclusions & Relevance: Based on this large cohort of children with CNLDO, probing between 9 and 15 months of age may be reasonable given that the rate of spontaneous resolution plateaued after 9 months and initial probing success declined after 15 months. This time frame supports both an earlier and narrower range of ages for intervention compared to the current practice of probing after one year of age.

Knowledge of systemic and ophthalmic manifestations combined with an understanding of the epidemiology of disease vectors is crucial for the diagnosis of tick-borne diseases.

We previously reported that the cellular transcription factor, HIF-1α, binds a hypoxia-response element (HRE) located within the promoter of Epstein-Barr virus's (EBV's) latent-lytic switch BZLF1 gene, Zp, inducing viral reactivation. Here, EBV-infected cell lines derived from gastric cancers and Burkitt lymphomas were incubated with HIF-1α-stabilizing drugs: iron chelator [Desferal® (DFO,)]; neddylation inhibitor [Pevonedistat® (MLN-4924)]; and prolyl hydroxylase inhibitor [Roxadustat® (FG-4592)]. DFO and MLN-4924, but not FG-4592, induced accumulation of both lytic EBV proteins and phosphorylated p53 in cell lines that contain a wild-type p53 gene. FG-4592 also failed to activate transcription from Zp in a reporter assay despite inducing accumulation of HIF-1α and transcription from another HRE-containing promoter. Unexpectedly, DFO failed to induce EBV reactivation in cell lines that express mutant or no p53 or when p53 expression was knocked down with shRNAs. Likewise, HIF-1α failed to activate transcription from Zp when p53 was knocked out by CRISPR-Cas9. Importantly, DFO induced binding of p53 as well as HIF-1α to Zp in ChIP assays, but only when the HRE was present. Nutlin-3, a drug known to induce accumulation of phosphorylated p53, synergized with DFO and MLN-4924 in inducing EBV reactivation. Conversely, KU-55933, a drug that inhibits ataxia telangiectasia mutated thereby preventing p53 phosphorylation, inhibited DFO-induced EBV reactivation. Lastly, activation of Zp transcription by DFO and MLN-4924 mapped to its HRE. Thus, we conclude that induction of BZLF1 gene expression by HIF-1α requires phosphorylated, wild-type p53 as a co-activator, with HIF-1α binding recruiting p53 to Zp. IMPORTANCE EBV, a human herpesvirus, is latently present in most nasopharyngeal carcinomas, Burkitt lymphomas, and some gastric cancers. To develop a lytic-induction therapy for treating patients with EBV-associated cancers, we need a way to efficiently reactivate EBV into lytic replication. EBV's BZLF1 gene product, Zta, usually controls this reactivation switch. We previously showed that HIF-1α binds the BZLF1 gene promoter, inducing Zta synthesis, and HIF-1α-stabilizing drugs can induce EBV reactivation. Here, we determined which EBV-positive cell lines are reactivated by classes of HIF-1α-stabilizing drugs. We found, unexpectedly, that HIF-1α-stabilizing drugs only induce reactivation when they also induce accumulation of phosphorylated, wild-type p53. Fortunately, p53 phosphorylation can also be provided by drugs such as Nutlin-3, leading to synergistic reactivation of EBV. These findings indicate that some HIF-1α-stabilizing drugs may be helpful as part of a lytic-induction therapy for treating patients with EBV-positive malignancies that contain wild-type p53.