Assessment of Two Novel Live-Attenuated Vaccine Candidates for Herpes Simplex Virus 2 (HSV-2) in Guinea Pigs

Vaccines

2021 Mar 13

Joyce, J;Patel, A;Murphy, B;Carr, D;Gershburg, E;Bertke, A;
| DOI: 10.3390/vaccines9030258

Treatment to ameliorate the symptoms of infection with herpes simplex virus 2 (HSV-2) and to suppress reactivation has been available for decades. However, a safe and effective preventative or therapeutic vaccine has eluded development. Two novel live-attenuated HSV-2 vaccine candidates (RVx201 and RVx202) have been tested preclinically for safety. Hartley guinea pigs were inoculated vaginally (n = 3) or intradermally (n = 16) with either vaccine candidate (2 × 107 PFU) and observed for disease for 28 days. All animals survived to study end without developing HSV-2-associated disease. Neither vaccine candidate established latency in dorsal root or sacral sympathetic ganglia, as determined by viral DNA quantification, LAT expression, or explant reactivation. Infectious virus was shed in vaginal secretions for three days following vaginal inoculation with RVx202, but not RVx201, although active or latent HSV-2 was not detected at study end. In contrast, guinea pigs inoculated with wild-type HSV-2 MS (2 × 105 PFU) vaginally (n = 5) or intradermally (n = 16) developed acute disease, neurological signs, shed virus in vaginal secretions, experienced periodic recurrences throughout the study period, and had latent HSV-2 in their dorsal root and sacral sympathetic ganglia at study end. Both vaccine candidates generated neutralizing antibody. Taken together, these findings suggest that these novel vaccine candidates are safe in guinea pigs and should be tested for efficacy as preventative and/or therapeutic anti-HSV-2 vaccines.

Dual targeting of brain region-specific kinases potentiates neurological rescue in Spinocerebellar ataxia type 1

The EMBO journal

2021 Mar 11

Lee, WS;Lavery, L;Rousseaux, MWC;Rutledge, EB;Jang, Y;Wan, YW;Wu, SR;Kim, W;Al-Ramahi, I;Rath, S;Adamski, CJ;Bondar, VV;Tewari, A;Soleimani, S;Mota, S;Yalamanchili, HK;Orr, HT;Liu, Z;Botas, J;Zoghbi, HY;
PMID: 33709453 | DOI: 10.15252/embj.2020106106

A critical question in neurodegeneration is why the accumulation of disease-driving proteins causes selective neuronal loss despite their brain-wide expression. In Spinocerebellar ataxia type 1 (SCA1), accumulation of polyglutamine-expanded Ataxin-1 (ATXN1) causes selective degeneration of cerebellar and brainstem neurons. Previous studies revealed that inhibiting Msk1 reduces phosphorylation of ATXN1 at S776 as well as its levels leading to improved cerebellar function. However, there are no regulators that modulate ATXN1 in the brainstem-the brain region whose pathology is most closely linked to premature death. To identify new regulators of ATXN1, we performed genetic screens and identified a transcription factor-kinase axis (ZBTB7B-RSK3) that regulates ATXN1 levels. Unlike MSK1, RSK3 is highly expressed in the human and mouse brainstems where it regulates Atxn1 by phosphorylating S776. Reducing Rsk3 rescues brainstem-associated pathologies and deficits, and lowering Rsk3 and Msk1 together improves cerebellar and brainstem function in an SCA1 mouse model. Our results demonstrate that selective vulnerability of brain regions in SCA1 is governed by region-specific regulators of ATXN1, and targeting multiple regulators could rescue multiple degenerating brain areas.

Serial transplantation unmasks galectin-9 contribution to tumor immune escape in the MB49 murine model

Scientific reports

2021 Mar 04

Baloche, V;Rivière, J;Tran, TBT;Gelin, A;Bawa, O;Signolle, N;Diop, MBK;Dessen, P;Beq, S;David, M;Busson, P;
PMID: 33664349 | DOI: 10.1038/s41598-021-84270-1

Mechanisms of tumor immune escape are quite diverse and require specific approaches for their exploration in syngeneic tumor models. In several human malignancies, galectin-9 (gal-9) is suspected to contribute to the immune escape. However, in contrast with what has been done for the infiltrating cells, the contribution of gal-9 produced by malignant cells has never been demonstrated in an animal model. Therefore, we derived isogenic clones-either positive or negative for gal-9-from the MB49 murine bladder carcinoma cell line. A progressive and consistent reduction of tumor growth was observed when gal-9-KO cells were subjected to serial transplantations into syngeneic mice. In contrast, tumor growth was unaffected during parallel serial transplantations into nude mice, thus linking tumor inhibition to the enhancement of the immune response against gal-9-KO tumors. This stronger immune response was at least in part explained by changing patterns of response to interferon-γ. One consistent change was a more abundant production of CXCL10, a major inflammatory factor whose production is often induced by interferon-γ. Overall, these observations demonstrate for the first time that serial transplantation into syngeneic mice can be a valuable experimental approach for the exploration of novel mechanisms of tumor immune escape.

Clonal expansion and activation of tissue-resident memory-like Th17 cells expressing GM-CSF in the lungs of severe COVID-19 patients

Science immunology

2021 Feb 23

Zhao, Y;Kilian, C;Turner, JE;Bosurgi, L;Roedl, K;Bartsch, P;Gnirck, AC;Cortesi, F;Schultheiß, C;Hellmig, M;Enk, LUB;Hausmann, F;Borchers, A;Wong, MN;Paust, HJ;Siracusa, F;Scheibel, N;Herrmann, M;Rosati, E;Bacher, P;Kylies, D;Jarczak, D;Lütgehetmann, M;Pfefferle, S;Steurer, S;Zur-Wiesch, JS;Puelles, VG;Sperhake, JP;Addo, MM;Lohse, AW;Binder, M;Huber, S;Huber, TB;Kluge, S;Bonn, S;Panzer, U;Gagliani, N;Krebs, CF;
PMID: 33622974 | DOI: 10.1126/sciimmunol.abf6692

Hyperinflammation contributes to lung injury and subsequent acute respiratory distress syndrome (ARDS) with high mortality in patients with severe coronavirus disease 2019 (COVID-19). To understand the underlying mechanisms involved in lung pathology, we investigated the role of the lung-specific immune response. We profiled immune cells in bronchoalveolar lavage fluid and blood collected from COVID-19 patients with severe disease and bacterial pneumonia patients not associated with viral infection. By tracking T cell clones across tissues, we identified clonally expanded tissue-resident memory-like Th17 cells (Trm17 cells) in the lungs even after viral clearance. These Trm17 cells were characterized by a a potentially pathogenic cytokine expression profile of IL17A and CSF2 (GM-CSF). Interactome analysis suggests that Trm17 cells can interact with lung macrophages and cytotoxic CD8+ T cells, which have been associated with disease severity and lung damage. High IL-17A and GM-CSF protein levels in the serum of COVID-19 patients were associated with a more severe clinical course. Collectively, our study suggests that pulmonary Trm17 cells are one potential orchestrator of the hyperinflammation in severe COVID-19.

Membranous nephropathy in a patient with coronavirus disease 2019 (COVID-19): A case report

Clinical nephrology. Case studies

2021 Feb 19

Miao, J;Fidler, ME;Nasr, SH;Larsen, CP;Zoghby, ZM;
PMID: 33633925 | DOI: 10.5414/CNCS110379

Though respiratory, immune, and coagulation systems are major targets of coronavirus disease 2019 (COVID-19), kidney dysfunction, presenting with acute kidney injury (AKI), is also common. Most AKI cases in COVID-19 manifest as acute tubular injury (ATI) in conjunction with multiorgan failure. While initial renal pathological findings were limited to acute tubular necrosis and collapsing glomerulopathy, a recent case series reported a larger spectrum of findings. Here, we report a case of membranous nephropathy (MN) in an 81-year-old Hispanic man with underlying chronic kidney disease (CKD) stage 3 who developed ATI in the setting of COVID-19. The patient was hospitalized for hypoxic respiratory failure in the setting of AKI stage 3 with serum creatinine 7.1 mg/dL 6 days after a positive-SARS-CoV-2 screening. He was found to have nephrotic range proteinuria, glycosuria (with normal serum glucose), anemia, and hypoalbuminemia. Kidney biopsy showed ATI and early MN. Workup for primary and secondary MN was unrevealing, and serum PLA2R antibody was negative. No viral particles were observed in podocytes. Although the MN could be incidental, this observation raises the question of whether SARS-CoV-2 infection can trigger or worsen an underlying MN from an exaggerated immune response associated with COVID-19.

Corticostriatal control of defense behavior in mice induced by auditory looming cues

Nature communications

2021 Feb 15

Li, Z;Wei, JX;Zhang, GW;Huang, JJ;Zingg, B;Wang, X;Tao, HW;Zhang, LI;
PMID: 33589613 | DOI: 10.1038/s41467-021-21248-7

Animals exhibit innate defense behaviors in response to approaching threats cued by the dynamics of sensory inputs of various modalities. The underlying neural circuits have been mostly studied in the visual system, but remain unclear for other modalities. Here, by utilizing sounds with increasing (vs. decreasing) loudness to mimic looming (vs. receding) objects, we find that looming sounds elicit stereotypical sequential defensive reactions: freezing followed by flight. Both behaviors require the activity of auditory cortex, in particular the sustained type of responses, but are differentially mediated by corticostriatal projections primarily innervating D2 neurons in the tail of the striatum and corticocollicular projections to the superior colliculus, respectively. The behavioral transition from freezing to flight can be attributed to the differential temporal dynamics of the striatal and collicular neurons in their responses to looming sound stimuli. Our results reveal an essential role of the striatum in the innate defense control.

The spatiotemporal expression of TERT and telomere repeat binding proteins in the postnatal mouse testes

Andrologia

2021 Feb 05

Kosebent, EG;Ozturk, S;
PMID: 33544428 | DOI: 10.1111/and.13976

Telomeres consist of repetitive DNA sequences and telomere-associated proteins. Telomeres located at the ends of eukaryotic chromosomes undergo shortening due to DNA replication, genotoxic factors and reactive oxygen species. The short telomeres are elongated by the enzyme telomerase expressed in the germ line, embryonic and stem cells. Telomerase is in the structure of ribonucleoprotein composed of telomerase reverse transcriptase (TERT), telomerase RNA component (Terc) and other components. Among telomere-associated proteins, telomeric repeat binding factor 1 (TRF1) and 2 (TRF2) exclusively bind to the double-stranded telomeric DNA to regulate its length. However, protection of telomeres 1 (POT1) interacts with the single-stranded telomeric DNA to protect from DNA damage response. Herein, we characterised the spatial and temporal expression of the TERT, TRF1, TRF2 and POT1 proteins in the postnatal mouse testes at the ages of 6, 8, 16, 20, 29, 32 and 88 days by using immunohistochemistry. Significant differences in the spatiotemporal expression patterns and levels of these proteins were determined in the postnatal testes (p < .05). These findings indicate that TERT and telomere repeat binding proteins seem to be required for maintaining the length and structural integrity of telomeres in the spermatogenic cells from newborn to adult terms.

Activation of Preoptic Arginine Vasopressin Neurons Induces Hyperthermia in Male Mice

Endocrinology

2021 Feb 01

Tabarean, IV;
PMID: 33249461 | DOI: 10.1210/endocr/bqaa217

Arginine vasopressin (AVP) is a neuropeptide acting as a neuromodulator in the brain and plays multiple roles, including a thermoregulatory one. However, the cellular mechanisms of action are not fully understood. Carried out are patch clamp recordings and calcium imaging combined with pharmacological tools and single-cell RT-PCR to dissect the signaling mechanisms activated by AVP. Optogenetics combined with patch-clamp recordings were used to determine the neurochemical nature of these neurons. Also used is telemetry combined with chemogenetics to study the effect of activation of AVP neurons in thermoregulatory mechanisms. This article reports that AVP neurons in the medial preoptic (MPO) area release GABA and display thermosensitive firing activity. Their optogenetic stimulation results in a decrease of the firing rates of MPO pituitary adenylate cyclase-activating polypeptide (PACAP) neurons. Local application of AVP potently modulates the synaptic inputs of PACAP neurons, by activating neuronal AVPr1a receptors and astrocytic AVPr1b receptors. Chemogenetic activation of MPO AVP neurons induces hyperthermia. Chemogenetic activation of all AVP neurons in the brain similarly induces hyperthermia and, in addition, decreases the endotoxin activated fever as well as the stress-induced hyperthermia.

Tobacco Hornworm (Manduca Sexta) Caterpillarhemolymph Modulates Reactive Oxygen Species and Calcium Generation in Tomato Protoplasts

Biophysical Journal

2021 Feb 01

Gandhi, A;Kariyat, R;Sahoo, N;
| DOI: 10.1016/j.bpj.2020.11.560

Plants have been at war with herbivorous insects for millions of years and have developed a set of highly regulated defense strategies to sense herbivore attack using chemical cues known as herbivore-associated elicitors (HAEs), including oral secretions, ovipositional fluids, and frass. These HAEs induce a series of signaling cascades, which ultimately provide induced defenses against them. Despite the existing HAEs and their role in plant defense induction, our knowledge of other herbivore generated HAEs in plant-herbivore interactions are limited. in this study, we demonstrate that ‘‘hemolymph’’ from Manduca sexta, caterpillarsalso induce plant defense signaling cascade and thereby act as an HAE.Using a dye-based imaging technique, our study showed that the application of crude M. sexta hemolymph potently increased Reactive Oxygen Species (ROS) productionin isolated tomato protoplasts. The addition of antioxidant NAC (N-acetyl-L-cysteine) antagonized hemolymph-induced ROS generation, indicating that M. sexta hemolymph is a ROS inducer in isolated protoplasts. Furthermore, incubating the protoplasts with Calcium (Ca2þ)chelator, BAPTA-AM efficiently abolished the hemolymph-induced ROS production, suggesting possible crosstalk between Ca2þ and ROS signaling. interestingly, the application of crude M. sexta hemolymph dramatically increased Ca2þ in tomato protoplasts. Also, hemolymph-mediated ROS and Ca2þ increase was inhibited in the absence of extracellular Ca2þ. Taken together, our study demonstrates that ‘‘hemolymph’’ from Manduca sexta can directly modulate intracellular ROS and Ca2þ production and possibly regulate defenses against insect herbivores by acting as an HAE.

Chloroquine diphosphate suppresses liver cancer via inducing apoptosis in Wistar rats using interventional therapy

Oncology Letters

2021 Jan 26

Hao, X;Li, W;
| DOI: 10.3892/ol.2021.12494

Liver cancer ranks as the second leading cause of cancer‑associated mortality worldwide. To date, neither current ablation therapy nor chemotherapy are considered ideal in improving the outcome of liver cancer. Therefore, more effective therapies for treating this devastating disease are urgently required. Interventional therapy has been used for numerous years in the treatment of different types of cancer, and is characterized by the direct delivery of anticancer drugs into the tumor. It has been reported that antimalarial chloroquine diphosphate (CQ) exerts effective anticancer activity against several types of cancer. However, its effect on liver cancer remains unclear. Therefore, in the present study, 2D monolayer cell culture and 3D spheroid _in vitro_ models, and a rat model, were utilized to investigate the effect of CQ on liver cancer. CQ demonstrated an effective anticancer effect on HepG2 cells and 3D liver spheroids. Furthermore, the drug significantly inhibited cell growth and viability in the 2D and 3D _in vitro_ models. The CQ‑based intervention treatment effectively attenuated tumor size and weight, increased food intake and consumption of drinking water, and improved body weight and survival rate of rats in the in vivo model. In addition, treatment with CQ potently increased the expression levels of the apoptosis‑related genes. Taken together, the findings of the present study may provide a novel insight into the development of safe and effective treatments for liver cancer.

Circuits between infected macrophages and T cells in SARS-CoV-2 pneumonia

Nature

2021 Jan 11

Grant, RA;Morales-Nebreda, L;Markov, NS;Swaminathan, S;Querrey, M;Guzman, ER;Abbott, DA;Donnelly, HK;Donayre, A;Goldberg, IA;Klug, ZM;Borkowski, N;Lu, Z;Kihshen, H;Politanska, Y;Sichizya, L;Kang, M;Shilatifard, A;Qi, C;Lomasney, JW;Argento, AC;Kruser, JM;Malsin, ES;Pickens, CO;Smith, SB;Walter, JM;Pawlowski, AE;Schneider, D;Nannapaneni, P;Abdala-Valencia, H;Bharat, A;Gottardi, CJ;Budinger, GRS;Misharin, AV;Singer, BD;Wunderink, RG;NU SCRIPT Study Investigators, ;
PMID: 33429418 | DOI: 10.1038/s41586-020-03148-w

Some patients infected with Severe Acute Respiratory Syndrome Coronavirus-2 (SARS-CoV-2) develop severe pneumonia and the acute respiratory distress syndrome (ARDS)1. Distinct clinical features in these patients have led to speculation that the immune response to virus in the SARS-CoV-2-infected alveolus differs from other types of pneumonia2. We collected bronchoalveolar lavage fluid samples from 88 patients with SARS-CoV-2-induced respiratory failure and 211 patients with known or suspected pneumonia from other pathogens and subjected them to flow cytometry and bulk transcriptomic profiling. We performed single-cell RNA-seq on 10 bronchoalveolar lavage fluid samples collected from patients with severe COVID-19 within 48 hours of intubation. In the majority of patients with SARS-CoV-2 infection, the alveolar space was persistently enriched in T cells and monocytes. Bulk and single-cell transcriptomic profiling suggested that SARS-CoV-2 infects alveolar macrophages, which in turn respond by producing T cell chemoattractants. These T cells produce interferon-gamma to induce inflammatory cytokine release from alveolar macrophages and further promote T cell activation. Collectively, our results suggest that SARS-CoV-2 causes a slowly unfolding, spatially limited alveolitis in which alveolar macrophages harboring SARS-CoV-2 and T cells form a positive feedback loop that drives persistent alveolar inflammation.

Endogenous Opioid Signaling in the Mouse Retina Modulates Pupillary Light Reflex

International journal of molecular sciences

2021 Jan 08

Cleymaet, AM;Berezin, CT;Vigh, J;
PMID: 33429857 | DOI: 10.3390/ijms22020554

Opioid peptides and their receptors are expressed in the mammalian retina; however, little is known about how they might affect visual processing. The melanopsin-expressing intrinsically photosensitive retinal ganglion cells (ipRGCs), which mediate important non-image-forming visual processes such as the pupillary light reflex (PLR), express β-endorphin-preferring, µ-opioid receptors (MORs). The objective of the present study was to elucidate if opioids, endogenous or exogenous, modulate pupillary light reflex (PLR) via MORs expressed by ipRGCs. MOR-selective agonist [D-Ala2, MePhe4, Gly-ol5]-enkephalin (DAMGO) or antagonist D-Phe-Cys-Tyr-D-Trp-Arg-Thr-Pen-Thr-NH2 (CTAP) was administered via intravitreal injection. PLR was recorded in response to light stimuli of various intensities. DAMGO eliminated PLR evoked by light with intensities below melanopsin activation threshold but not that evoked by bright blue irradiance that activated melanopsin signaling, although in the latter case, DAMGO markedly slowed pupil constriction. CTAP or genetic ablation of MORs in ipRGCs slightly enhanced dim-light-evoked PLR but not that evoked by a bright blue stimulus. Our results suggest that endogenous opioid signaling in the retina contributes to the regulation of PLR. The slowing of bright light-evoked PLR by DAMGO is consistent with the observation that systemically applied opioids accumulate in the vitreous and that patients receiving chronic opioid treatment have slow PLR.

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	Description
sense Example: Hs-LAG3-sense	Standard probes for RNA detection are in antisense. Sense probe is reverse complent to the corresponding antisense probe.
Intron# Example: Mm-Htt-intron2	Probe targets the indicated intron in the target gene, commonly used for pre-mRNA detection
Pool/Pan Example: Hs-CD3-pool (Hs-CD3D, Hs-CD3E, Hs-CD3G)	A mixture of multiple probe sets targeting multiple genes or transcripts
No-XSp Example: Hs-PDGFB-No-XMm	Does not cross detect with the species (Sp)
XSp Example: Rn-Pde9a-XMm	designed to cross detect with the species (Sp)
O# Example: Mm-Islr-O1	Alternative design targeting different regions of the same transcript or isoforms
CDS Example: Hs-SLC31A-CDS	Probe targets the protein-coding sequence only
EnEm	Probe targets exons n and m
En-Em	Probe targets region from exon n to exon m
Retired Nomenclature
tvn Example: Hs-LEPR-tv1	Designed to target transcript variant n
ORF Example: Hs-ACVRL1-ORF	Probe targets open reading frame
UTR Example: Hs-HTT-UTR-C3	Probe targets the untranslated region (non-protein-coding region) only
5UTR Example: Hs-GNRHR-5UTR	Probe targets the 5' untranslated region only
3UTR Example: Rn-Npy1r-3UTR	Probe targets the 3' untranslated region only
Pan Example: Pool	A mixture of multiple probe sets targeting multiple genes or transcripts

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