Catch me if you can – the crosstalk of ZIKV and the restriction factor Tetherin

Zika virus (ZIKV) is a flavivirus that is mainly transmitted by Aedes mosquitos and normally causes mild symptoms. During the outbreak in the Americas in 2015, it was associated with more severe implications, like microcephaly in new-borns and the Gullain-Barré syndrome. The lack of specific vaccines and cures strengthen the need for a deeper understanding of the virus life cycle and virus-host interactions. The restriction factor tetherin (THN) is an interferon-inducible cellular protein with broad antiviral properties. It is known to inhibit the release of various enveloped viruses by tethering them to each other and to the cell membrane, thereby preventing their further spread. On the other hand, different viruses have developed various escape strategies against THN. Analysis of the crosstalk between ZIKV and THN revealed that in spite of a strong induction of THN mRNA expression in ZIKV-infected cells, this is not reflected by an elevated protein level of THN. Contrariwise, the THN protein level is decreased due to a reduced half-life. The increased degradation of THN in ZIKV infected cells involves the endo-lysosomal system, but does not depend on the early steps of autophagy. Enrichment of THN by depletion of the ESCRT-0 protein HRS diminishes ZIKV release and spread, which points out the capacity of THN to restrict ZIKV and explains the enhanced THN degradation in infected cells as an effective viral escape strategy. Importance Although tetherin expression is strongly induced by ZIKV infection there is a reduction in the amount of tetherin protein. This is due to an enhanced lysosomal degradation. However, if tetherin level is rescued release of ZIKV is impaired. This shows that tetherin is a restriction factor for ZIKV and the induction of an efficient degradation represents a viral escape strategy. To our knowledge this is the first study that describes and characterizes tetherin as an restriction factor for ZIKV life cycle.

A Case of Trimethoprim-Sulfamethoxazole-Induced Aseptic Meningitis Masquerading as Septic Shock

ABSTRACT Trimethoprim-sulfamethoxazole-induced aseptic meningitis (TSIAM) is a rare adverse reaction to a commonly prescribed antibiotic. We describe a case of severe TSIAM which resembled septic shock. A 30-year-old male with relapsed Hodgkin’s lymphoma 25 days status post autologous stem cell transplant presented to our clinic for evaluation of trimethoprim-sulfamethoxazole (TMP-SMX) hypersensitivity. After review of patient’s history and records, we had a low suspicion for a TMP-SMX adverse reaction and conducted an oral challenge to one 160 mg/800 mg tab of TMP-SMX. Four hours later, the patient developed vomiting, lightheadedness, and disorientation with progression to rigors, fever, tachycardia, and hypotension. He was admitted for fluid resuscitation and broad-spectrum antibiotic coverage for neutropenic fever and possible septic shock. A lumbar puncture performed due to complaints of headache, photophobia, and neck pain showed 375 white blood cells/µL with 73% neutrophil predominance, normal glucose (75 mg/dL), and elevated protein (101 mg/dL); additional cerebrospinal fluid (CSF) studies were negative for infectious etiologies. Fever and headache resolved by hospital day 4, at which time patient was discharged home. We believe this case represents TSIAM given the characteristic timing of symptom onset, CSF findings, and timing of symptom resolution without other clear etiology found on extensive infectious evaluation. It is important for allergists to recognize TSIAM, including its potential presentation as shock, in order to appropriately diagnose and counsel patients who seek evaluation for TMP-SMX adverse reactions.

Mechanism of Action of Bone Marrow Mesenchymal Stem Cell in Combination with miR-36b Therapy in Cellular Repair of Septic Lung Injury

Our study aims to assess the role of BMSCs (MSCs) transplantation in combination with miR-36b in the repair of septic lung injury. MSCs were cultured by the paste-wall method and characterized. MSCs combined with miR-36b medium were added to lung-injured cells for 14 days followed by analysis of cell viability by CCK-8 assay, GLUT3 expression and apoptosis by western blot. After 1 and 3 days of growth of MSCs progeny under electron microscopy, the MSCs showed long shuttle-shaped morphology. MSCs in combination with miR-36b resulted in enhanced proliferative capacity of lung-injured cells and enhanced protein expression of GLUT3. CCK-8 assay showed increased viability of lung-injured cells and elevated protein and mRNA expression of GLUT3. Meanwhile, the expression of apoptosis-related proteins was significantly down-regulated. In conclusion, MSCs in combination with miR-36b therapy may ameliorate lung injury by promoting lung cell proliferation through inhibition of apoptotic pathway.

Epithelioid haemangioendothelioma – A rare cause of pleural effusion

We present the case of a 44-year-old patient, with a history of lymphatic tuberculosis, diagnosed with a pleural effusion associated with pulmonary consolidations. Pleural effusions are frequent in the general population, presenting an incidence of 3–4 cases/1000 people. The etiological diagnosis can be difficult, as they can be secondary to a variety of diseases. Numerous extrapulmonary disorders can explain fluid accumulations in the pleural space with elevated protein level (exudates). However, more frequently they are the result of pulmonary diseases – inflammatory, infectious or neoplastic. The case entailed a differential diagnosis between pulmonary and pleural tuberculosis and neoplastic diseases. Despite receiving antituberculous treatment, the patient’s initial symptoms worsen, the patient starts presenting new signs and symptoms, and the fluid’s characteristics change – expression of an unfavourable evolution. Establishing the diagnosis proved to be difficult as invasive procedures were necessary. We diagnosed an extremely rare tumour, of vascular origin as the cause of the pleural effusion. Epithelioid haemangioendothelioma (EHE) presents an incidence of 1 in 1 million people. In conclusion, when treating a patient for pleural effusion, the lack of improvement entails reconsidering the initial diagnosis and performing more extensive tests.

QKI Promotes Hypoxia Induced Smooth Muscle Reprogramming in Pulmonary Hypertension

Background: Pulmonary hypertension (PH) is a complex and progressive cardiopulmonary disorder with poor prognosis and limited therapeutic treatments. Recent evidence suggests that RNA binding proteins (RBPs) participate in the pathogenesis of human and experimental pulmonary arterial hypertension. Quaking (QKI) as an important RBPs is involved in mRNA biogenesis, export, decay and translation. However, the biological significance of QKI in phenotypic transformation of PASMCs in PH as well as in abnormal pulmonary vascular remodeling remain elusive. Methods: We assessed the expression pattern, phenotypic transformation effect, and mechanism of QKI in rodent Su/Hx-induced PH model, Human PAH samples and in HPASMCs.Results: Elevated protein expression level of QKI was found in animal PH and human PAH samples, thus in hypoxic HPASMCs. Inhibition of QKI attenuated proliferation and phenotype switching in HPASMCs. Mechanistically, QKI was found to mediate STAT3 mRNA stabilization by binding to its 3’Untranslated Region (3’-UTR). Downregulation of QKI attenuated STAT3 expression in PASMCs, while overexpression of STAT3 in PASMCs was widely regarded to be involved in the progression of PH. In addition, as a transcription factor, STAT3 was identified to bound to miR-146b promoter to induce its expression, while miR-146b was proved to promote smooth muscle reprogramming through inhibiting STAT1 and TET2 expression during pulmonary vascular remodeling.Conclusions: Our study demonstrates the QKI-STAT3-miR-146b pathway as a novel mechanistic insights into hypoxic reprogramming that permits vascular remodeling, and thus provides proof of concept for anti-remodeling therapy through the direct modulation this axis in PH.

CTIM-17. INTRA-CRANIAL ADMINISTRATION OF CTLA-4 AND PD-1 IMMUNE CHECKPOINT-INHIBITING MONOCLONAL ANTIBODIES IN RECURRENT GLIOBLASTOMA PATIENTS: A MULTI-COHORT ADAPTIVE PHASE I CLINICAL TRIAL

BACKGROUND: Intracerebral (iCE) administration of nivolumab (NIVO) and ipilimumab (IPI) after resection of recurrent glioblastoma (rGB), followed by repeated intravenous(IV) NIVO was recently shown to be feasible, safe and associated with encouraging survival. Subsequent cohorts were defined to investigate the addition of biweekly intracavitary (iCA) or intrathecal (iTH) NIVO +/- IPI administrations. METHODS Four groups were defined according to rGB resectability and postoperative treatment schedule. Group A and D underwent biopsy, B and C maximal safe resection. All patients received iCE injections of 10 mg/1ml NIVO + 5 mg/1ml IPI at the end of surgery, after which an Ommaya catheter was implanted iCA (A, B and C) or iTH (D). Following surgery, all patients received biweekly IV low-dose NIVO(10mg) combined with iCA/iTH 10 mg NIVO (A and B) + 1, 5 or 10 mg IPI (C and D) for up to 24 weeks. NIVO/IPI concentrations were dosed in the cerebrospinal fluid (CSF). Gene sequencing and RNA gene expression profiling were performed on all tissue samples RESULTS 39pts(27 male; 16 in A, 16 in B, 4 in C, 3 in D; recruitment ongoing in C+D) were enrolled. All patients received the predefined dose of iCE IPI/NIVO. Most frequent AEs were fatigue (n=30), headache (n=19), confusion (n=14), dysphasia (n=13), and fever (n=10). Ommaya infection occurred in 5patients, subacute neurological deterioration requiring corticosteroids in 6patients. There were no G5 AEs. irAEs were infrequent and mild. Median PFS and OS were 5w(95% CI 1-8) and 23w(95% CI 0-53) in A and 13w(95% CI 7-19) and 42w(95% CI 30-54) in B, respectively. >90% of CSF samples had elevated protein levels and lymphocytic pleocytosis. There was no evidence for accumulation of NIVO/IPI in the CSF. CONCLUSION Repeated intracavitary or intrathecal administration of NIVO +/- IPI in rGB is feasible and safe. Favourable survival outcome is seen in patients amenable to surgical resection.

Protease-controlled secretion and display of intercellular signals

To program intercellular communication for biomedicine, it is crucial to regulate the secretion and surface display of signaling proteins. If such regulations are at the protein level, there are additional advantages, including compact delivery and direct interactions with endogenous signalling pathways. We created a modular, generalizable design called Retained Endoplasmic Cleavable Secretion (RELEASE), with engineered proteins retained in the endoplasmic reticulum and displayed/secreted in response to specific proteases. The design allows functional regulation of multiple synthetic and natural proteins by synthetic protease circuits to realize diverse signal processing capabilities, including logic operation and threshold tuning. By linking RELEASE to additional novel sensing and processing circuits, we were able to achieve elevated protein secretion in response to undruggable oncogene KRAS mutants. RELEASE should enable the local, programmable delivery of intercellular cues for a broad variety of fields such as neurobiology, cancer immunotherapy and cell transplantation.

Next-generation sequencing of cerebrospinal fluid for diagnosis of atypical herpes simplex encephalitis

Objectives Herpes simplex encephalitis (HSE) is one of the most common causes of severe viral encephalitis. The characteristic manifestations of HSE include cerebrospinal fluid with mild cytopenia, dominated by lymphocytes, elevated protein, and normal blood glucose values (3.9–6.1 mmol/L). Although it is not difficult to diagnose classical HSE, diagnosing clinically atypical cases is more difficult. Methods We reviewed the results of next-generation sequencing (NGS) of CSF in a series of patients diagnosed with atypical HSE. Results Four patients lacking classical clinical manifestations of HSE, including no fever, headache, or other typical neurological deficit symptoms, 1–2 × 106 cells/L CSF leucocyte count, and no typical imaging features, were diagnosed with atypical HSE by NGS of CSF. The NGS reads corresponding to herpes simplex virus type 1 ranged from 2 to 13,174. Conclusions Mild HSE may not present with classic frontotemporal lobe syndrome and fever may not be an inevitable symptom in patients with immunosuppression. However, the possibility of HSE should be considered in patients with atypical intracranial infection, and these patients should be tested by NGS.

Autoimmune glial fibrillary acidic protein astrocytopathy presented as ataxia, myoclonus and bulbar syndrome: a case report and review of the literature

ObjectiveTo describe an atypical case of autoimmune glial fibrillary acidic protein (GFAP) astrocytopathy.ResultsA patient in his 60s presented with 6 months of progressive ataxia, proximal myoclonus and bulbar symptomatology. Cerebrospinal fluid (CSF) analysis showed monocytic pleocytosis, elevated protein level and elevated adenosine deaminase (ADA) level. CSF microbiological studies were negative and brain and cervical MRI showed no significant findings. We tested for nuclear, cytoplasmatic and synaptic neural autoantibodies as well as anti-GFAP antibodies. While awaiting these results, the patient was commenced on methylprednisolone boluses (1 g/day for 5 days), noting rapid neurological improvement. Eventually, CSF tests were positive for anti-GFAP antibodies.ConclusionWe report atypical manifestations of GFAP astrocytopathy. Further research is needed to fully understand the spectrum of neurological manifestations of this autoimmune disease and facilitate timely diagnosis.