Probes for INS

Regulation of translation is essential for the diverse biological processes involved in development. Particularly, mammalian oocyte development requires the precisely controlled translation of maternal transcripts to coordinate meiotic and early embryo progression while transcription is silent. It has been recently reported that key components of mRNA translation control are short and long noncoding RNAs (ncRNAs). We found that the ncRNA Brain cytoplasmic 1 (BC1) has a role in the fully grown germinal vesicle (GV) mouse oocyte, where is highly expressed in the cytoplasm associated with polysomes. Overexpression of BC1 in GV oocyte leads to a minute decrease in global translation with a significant reduction of specific mRNA translation via interaction with the Fragile X Mental Retardation Protein (FMRP). BC1 performs a repressive role in translation only in the GV stage oocyte without forming FMRP or Poly(A) granules. In conclusion, BC1 acts as the translational repressor of specific mRNAs in the GV stage via its binding to a subset of mRNAs and physical interaction with FMRP. The results reported herein contribute to the understanding of the molecular mechanisms of developmental events connected with maternal mRNA translation.

Angiotensin-converting enzyme (ACE), a key element of the renin-angiotensin system (RAS), has recently been identified as a new marker of both adult and embryonic human hematopoietic stem/progenitor cells (HSPCs). However, whether a full renin-angiotensin pathway is locally present during the hematopoietic emergence is still an open question. In the present study, we show that this enzyme is expressed by hematopoietic progenitors in the developing mouse embryo. Furthermore, ACE and the other elements of RAS-namely angiotensinogen, renin, and angiotensin II type 1 (AT1) and type 2 (AT2) receptors-are expressed in the paraaortic splanchnopleura (P-Sp) and in its derivative, the aorta-gonad-mesonephros region, both in human and mouse embryos. Their localization is compatible with the existence of a local autocrine and/or paracrine RAS in these hemogenic sites. In vitro perturbation of the RAS by administration of a specific AT1 receptor antagonist inhibits almost totally the generation of blood CD45-positive cells from dissected P-Sp, implying that angiotensin II signaling is necessary for the emergence of hematopoietic cells. Conversely, addition of exogenous angiotensin II peptide stimulates hematopoiesis in culture, with an increase in the number of immature c-Kit+ CD41+ CD31+ CD45+ hematopoietic progenitors, compared to the control. These results highlight a novel role of local-RAS during embryogenesis, suggesting that angiotensin II, via activation of AT1 receptor, promotes the emergence of undifferentiated hematopoietic progenitors.

Mutations of Odontogenesis-Associated Phosphoprotein (ODAPH, OMIM *614829) cause autosomal recessive amelogenesis imperfecta, however, the function of ODAPH during amelogenesis is unknown. Here we characterized normal Odaph expression by in situ hybridization, generated Odaph truncation mice using CRISPR/Cas9 to replace the TGC codon encoding Cys41 into a TGA translation termination codon, and characterized and compared molar and incisor tooth formation in Odaph+/+, Odaph+/C41*, and OdaphC41*/C41* mice. We also searched genomes to determine when Odaph first appeared phylogenetically. We determined that tooth development in Odaph+/+ and Odaph+/C41* mice was indistinguishable in all respects, so the condition in mice is inherited in a recessive pattern, as it is in humans. Odaph is specifically expressed by ameloblasts starting with the onset of post-secretory transition and continues until mid-maturation. Based upon histological and ultrastructural analyses, we determined that the secretory stage of amelogenesis is not affected in OdaphC41*/C41* mice. The enamel layer achieves a normal shape and contour, normal thickness, and normal rod decussation. The fundamental problem in OdaphC41*/C41* mice starts during post-secretory transition, which fails to generate maturation stage ameloblasts. At the onset of what should be enamel maturation, a cyst forms that separates flattened ameloblasts from the enamel surface. The maturation stage fails completely.

In mammalian testes, undifferentiated spermatogonia (Aundiff) undergo differentiation in response to retinoic acid (RA), while their progenitor states are partially maintained by fibroblast growth factors (FGFs). Sertoli valve (SV) is a region located at the terminal end of seminiferous tubule (ST) adjacent to the rete testis (RT), where the high density of Aundiff is constitutively maintained with the absence of active spermatogenesis. However, the molecular and cellular characteristics of SV epithelia still remain unclear. In this study, we first identified the region-specific AKT phosphorylation in the SV Sertoli cells and demonstrated non-cell autonomous specialization of Sertoli cells in the SV region by performing a Sertoli cell ablation/replacement experiment. The expression of Fgf9 was detected in the RT epithelia, while the exogenous administration of FGF9 caused ectopic AKT phosphorylation in the Sertoli cells of convoluted ST. Furthermore, we revealed the SV region-specific expression of Cyp26a1, which encodes an RA-degrading enzyme, and demonstrated that the increased RA levels in the SV region disrupt its pool of Aundiff by inducing their differentiation. Taken together, RT-derived FGFs and low levels of RA signaling contribute to the non-cell-autonomous regionalization of the SV epithelia and its local maintenance of Aundiff in the SV region.

We investigate as yet an unidentified role of NOX1, a non-phagocytic isoform of the superoxide-generating NADPH oxidase, in immune responses using Nox1-knockout mice (Nox1-KO). The transcripts of NOX1 was expressed in lymphoid tissues, including the spleen, thymus, bone marrow, and inguinal lymphoid nodes. When antibody production after ovalbumin (OVA) immunization was examined, no significant differences were observed in serum anti-OVA IgG levels between wild-type mice (WT) and Nox1-KO. In the experimental asthma, the infiltration of eosinophils and the Th2 cytokine response after the induction of asthma with OVA were similar between the two genotypes. However, the severity and incidence of experimental collagen-induced arthritis (CIA) following the administration of a low dose of endotoxin (LPS) were significantly lower in Nox1-KO. While neither serum levels of autoantibodies nor in vitro cytokine responses were affected by Nox1 deficiency, NOX1 mRNA levels in the spleen significantly increased after the LPS challenge. Among the spleen cells, remarkable LPS-induced upregulation of NOX1 was demonstrated in both CD11b+ monocytes/macrophages and CD11c+ dendritic cells, suggesting that LPS-inducible NOX1 in monocytes/macrophages/dendritic cells may modulate the development of experimental CIA. Therapeutic targeting of NOX1 may therefore control the onset and/or severity of arthritis which is exacerbated by bacterial infection.

Opioid signaling has been shown to be critically important in the neuromodulation of sensory circuits in the superficial spinal cord. Agonists of the mu-opioid receptor (MOR) elicit itch, whereas agonists of the kappa-opioid receptor (KOR) have been shown to inhibit itch. Despite the clear roles of MOR and KOR for the modulation itch, whether the delta-opioid receptor (DOR) is involved in the regulation of itch remained unknown. Here, we show that intrathecal administration of DOR agonists suppresses chemical itch and that intrathecal application of DOR antagonists is sufficient to evoke itch. We identify that spinal enkephalin neurons co-express neuropeptide Y (NPY), a peptide previously implicated in the inhibition of itch. In the spinal cord, DOR overlapped with both the NPY receptor (NPY1R) and KOR, suggesting that DOR neurons represent a site for convergent itch information in the dorsal horn. Lastly, we found that neurons co-expressing DOR and KOR showed significant Fos induction following pruritogen-evoked itch. These results uncover a role for DOR in the modulation of itch in the superficial dorsal horn. Perspective: This article reveals the role of the delta-opioid receptor in itch. Intrathecal administration of delta agonists suppresses itch whereas the administration of delta antagonists is sufficient to induce itch. These studies highlight the importance of delta-opioid signaling for the modulation of itch behaviors, which may represent new targets for the management of itch disorders.

Growth hormone secretagogue receptor 1a (GHSR1a)-the receptor for orexigenic hormone ghrelin-is a G protein-coupled receptor that is widely distributed in the brain, including the hippocampus. Studies have demonstrated that genetic deletion of GHSR1a affects memory, suggesting the importance of ghrelin/GHSR1a signaling in cognitive control. However, current reports are controversial, and the mechanism underlying GHSR1a modulation of memory is uncertain. Here, we first report that global GHSR1a knockout enhances hippocampus-dependent memory, facilitates initial LTP in dorsal hippocampal Schaffer Collateral-CA1 synapses, and downregulates Akt activity in the hippocampus. Moreover, we show that the intrinsic excitability of GAD67+ interneurons-rather than neighboring pyramidal neurons in the dCA1-is suppressed by GHSR1a deletion, an effect that is antagonized by acute application of the Akt activator SC79. In addition, the inhibitory postsynaptic currents (IPSCs) on dCA1 pyramidal neurons are selectively reduced in mice with a GHSR1a deficiency. Finally, we demonstrate that selectively increasing the excitability of parvalbumin-expressing interneurons by hM3Dq-DREADDs increases IPSCs on dCA1 pyramidal neurons and normalizes memory in Ghsr1a KO mice. Our findings thus reveal a novel mechanism underlying memory enhancement of GHSR1a deficiency and herein support an adverse effect of GHSR1a signaling in hippocampus-dependent memory processes.

The COVID-19 pandemic continues to be a worldwide threat and effective antiviral drugs and vaccines are being developed in a joint global effort. However, some elderly and immune-compromised populations are unable to raise an effective immune response against traditional vaccines.We hypothesised that passive immunity engineered by the in vivo expression of anti-SARS-CoV-2 monoclonal antibodies (mAbs), an approach termed vectored-immunoprophylaxis (VIP), could offer sustained protection against COVID-19 in all populations irrespective of their immune status or age.We developed three key reagents to evaluate VIP for SARS-CoV-2: (i) we engineered standard laboratory mice to express human ACE2 via rAAV9 in vivo gene transfer, to allow in vivo assessment of SARS-CoV-2 infection, (ii) to simplify in vivo challenge studies, we generated SARS-CoV-2 Spike protein pseudotyped lentiviral vectors as a simple mimic of authentic SARS-CoV-2 that could be used under standard laboratory containment conditions and (iii) we developed in vivo gene transfer vectors to express anti-SARS-CoV-2 mAbs.A single intranasal dose of rAAV9 or rSIV.F/HN vectors expressing anti-SARS-CoV-2 mAbs significantly reduced SARS-CoV-2 mimic infection in the lower respiratory tract of hACE2-expressing mice. If translated, the VIP approach could potentially offer a highly effective, long-term protection against COVID-19 for highly vulnerable populations; especially immune-deficient/senescent individuals, who fail to respond to conventional SARS-CoV-2 vaccines. The in vivo expression of multiple anti-SARS-CoV-2 mAbs could enhance protection and prevent rapid mutational escape.

Human papillomavirus (HPV)-related multiphenotypic sinonasal carcinoma (HMSC) is newly suggested and characterized by HPV-related tumors. HMSC has a relatively good prognosis. No cases of brain invasion have been reported to date. We encountered a case of brain invasion by HMSC, in which we assessed the effectiveness of radiotherapy in comparison with biopsy and autopsy. A 69-year-old man was referred to a hospital three months after intracerebral hemorrhage (ICH). Contrast magnetic resonance imaging revealed a tumor in the ethmoid sinus involving the brain. We performed transnasal biopsy and intensity-modulated radiotherapy for sinonasal and intracranial lesions. Despite radiotherapy, the patient died on day 41 after radiation. Biopsy specimens displayed mixed findings of epithelial and mesenchymal components. The tumor was immunoreactive for p16, and the RNA in situ hybridization for HPV was positive. Finally, we diagnosed the patient as having HMSC. Autopsy of the sinonasal tissue revealed a reduction in the number of tumor cells. There was a marked reduction in the number of tumor cells in the sinonasal tissue compared to that in the invaded brain tissue. The effectiveness of radiotherapy could depend on the histopathological components and location of the lesion, even in the same patient.

Previous studies suggest that microglial-expressed Apolipoprotein E (ApoE) is necessary to shift microglia into a neurodegenerative transcriptional state in Alzheimer's disease (AD) mouse models. On the other hand, elimination of microglia shifts amyloid beta (Aβ) accumulation from parenchymal plaques to cerebral amyloid angiopathy (CAA), mimicking the effects of global APOE*4 knock-in. Here, we specifically knock-out microglial-expressed ApoE while keeping astrocytic-expressed ApoE intact. When microglial-specific ApoE is knocked-out of a 5xFAD mouse model of AD, we found a ~35% increase in average Aβ plaque size, but no changes in plaque load, microglial number, microglial clustering around Aβ plaques, nor the formation of CAA. Immunostaining revealed ApoE protein present in plaque-associated microglia in 5xFAD mice with microglial-specific ApoE knockout, suggesting that microglia can take up ApoE from other cellular sources. Mice with Apoe knocked-out of microglia had lower synaptic protein levels than control mice, indicating that microglial-expressed ApoE may have a role in synapse maintenance. Surprisingly, microglial-specific ApoE knock-out resulted in few differentially expressed genes in both 5xFAD and control mice; however, some rescue of 5xFAD associated neuronal networks may occur with microglial-specific ApoE knock-out as shown by weighted gene co-expression analysis. Altogether, our data indicates that microglial-expressed ApoE may not be necessary for plaque formation or for the microglial transcriptional shift into a Disease Associated Microglia state that is associated with reactivity to plaques but may be necessary for plaque homeostasis in disease and synaptic maintenance under normal conditions.

Chimeric antigen receptor (CAR) T cells have largely been successful in treating hematological malignancies in the clinic but have not been as effective in treating solid tumors, in part, owing to poor access and the immunosuppressive tumor microenvironment. In addition, CAR-T therapy can cause potentially life-threatening side effects, including cytokine release syndrome and neurotoxicity. Current preclinical testing of CAR-T therapy efficacy is typically performed in mouse tumor models, which often fails to predict toxicity. Recent developments in humanized models and transgenic mice as well as in vitro three-dimensional organoids in early development and nonhuman primate models are being adopted for CAR-T cell efficacy and toxicity assessment. However, because no single model perfectly recapitulates the human immune system and tumor microenvironment, careful model selection based on their respective pros and cons is crucial for adequate evaluation of different CAR-T treatments, so that their clinical development can be better supported.

PICO question In cats infected with papillomavirus, is the risk of developing feline cutaneous squamous cell carcinoma greater than cats that are not infected with papillomavirus?   Clinical bottom line Category of research question Risk The number and type of study designs reviewed Eleven papers were critically reviewed, nine were case-control studies and two were experimental in vitro studies Strength of evidence Moderate Outcomes reported Infection of feline epithelial skin cells with Felis catus papillomavirus type 2 (FcaPV-2) is a risk factor for the development of feline cutaneous squamous cell carcinoma. The pathogenesis of FcaPV-2 infection and neoplastic transformation into malignant cells shares similar pathways to the human papillomavirus (HPV) model of pathogenesis and carcinogenesis with some differences Conclusion In conclusion, there is moderate strength of evidence in the literature to support a role of FcaPV-2 in the development of cutaneous squamous cell carcinomas in cats. Therefore, prevention of infection with FcaPV-2 should prevent some cancers   How to apply this evidence in practice The application of evidence into practice should take into account multiple factors, not limited to: individual clinical expertise, patient’s circumstances and owners’ values, country, location or clinic where you work, the individual case in front of you, the availability of therapies and resources. Knowledge Summaries are a resource to help reinforce or inform decision making. They do not override the responsibility or judgement of the practitioner to do what is best for the animal in their care.