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Probes for INS

ACD can configure probes for the various manual and automated assays for INS for RNAscope Assay, or for Basescope Assay compatible for your species of interest.

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Circular Rims2 Deficiency Causes Retinal Degeneration

Advanced biology

2021 Nov 05

Sun, LF;Ma, Y;Ji, YY;Wu, Z;Wang, YH;Mou, H;Jin, ZB;
PMID: 34738746 | DOI: 10.1002/adbi.202100906

Circular RNAs (circRNAs) refer to a newly recognized family of non-coding RNA with single-stranded RNAs. Despite emerging evidence indicating that circRNAs are abundantly expressed in various tissues, especially in the brain and retina, the role of circRNAs in retinal function and diseases is still largely unknown. Circular Rims2 (circRims2) is highly expressed and conserved in both the human and mouse brains. However, little is known about the expression and function of circRims2 in the retina. In the current study, the high-throughput RNA-seq analysis reveals a high expression of circRims2 in the retina. In addition, it is found that circRims2 is mainly located in plexiform layers that contain synapses between retinal neurons. Knocking down circRims2 with short hairpin RNA through subretinal adeno-associated viral (AAV) delivery in the mice leads to the decrease of the thickness of the outer and inner segment (OS/IS) layers and outer nuclear layer (ONL), and cessation of scotopic and photopic electroretinogram responses. Furthermore, the current study finds that circRims2 deficiency evokes retinal inflammation and activates the tumor necrosis factor (TNF) signaling pathway. Therefore, circRims2 may play an important role in the maintenance of retinal structure and function, and circRims2 deficiency may lead to pathogenic changes in the retina.
NEAT1 在小鼠早期妊娠子宫组织中的表达和调节

中国畜牧兽医

2021 Jan 01

张新艳, ;贾燕妮, ;蔡芮, ;张瑞雪, ;靳亚平, ;林鹏飞, ;
| DOI: 10.16431/j.cnki.1671-7236.2021.11.016

The test was aimed to study the expression and regulation of nuclear paraspeckle assembly transcript 1 (NEAT1) in uterine tissue during early pregnancy of mice, and provided scientific evidence for revealing the mechanism of NEAT1 in mouse embryo implantation. The models of early pregnancy, pseudopregnancy, artificial decidualization and steroid hormone treatments were constructed respectively, and the expression of NEAT1 in the uterine tissue was detected by RNAscope and Real-time quantitative PCR techniques. The results showed that the expression of NEAT1 was dynamically changing in uterine tissue in early pregnancy. NEAT1 was detected in endometrium and muscular layer on days 1-5 of pregnancy and mainly observed in endometrial luminal and glandular epithelial cells. NEAT1 was highly expressed in decidual cells on days 6-8 of pregnancy. There was no detectable NEAT1 staining at conceptus. Similar with those of early pregnancy, the expression of NEAT1 in pseudopregnant uteri was observed in endometrium and muscular layer. In the artificial decidulization model, NEAT1 was mainly localized in the decidualised cells and the expression level was extremely significant higher than that on the control group (_P_
The Bank Vole (Clethrionomys glareolus)-Small Animal Model for Hepacivirus Infection

Viruses

2021 Dec 03

Röhrs, S;Begeman, L;Straub, BK;Boadella, M;Hanke, D;Wernike, K;Drewes, S;Hoffmann, B;Keller, M;Drexler, JF;Drosten, C;Höper, D;Kuiken, T;Ulrich, RG;Beer, M;
PMID: 34960690 | DOI: 10.3390/v13122421

Many people worldwide suffer from hepatitis C virus (HCV) infection, which is frequently persistent. The lack of efficient vaccines against HCV and the unavailability of or limited compliance with existing antiviral therapies is problematic for health care systems worldwide. Improved small animal models would support further hepacivirus research, including development of vaccines and novel antivirals. The recent discovery of several mammalian hepaciviruses may facilitate such research. In this study, we demonstrated that bank voles (Clethrionomys glareolus) were susceptible to bank vole-associated Hepacivirus F and Hepacivirus J strains, based on the detection of hepaciviral RNA in 52 of 55 experimentally inoculated voles. In contrast, interferon α/β receptor deficient C57/Bl6 mice were resistant to infection with both bank vole hepaciviruses (BvHVs). The highest viral genome loads in infected voles were detected in the liver, and viral RNA was visualized by in situ hybridization in hepatocytes, confirming a marked hepatotropism. Furthermore, liver lesions in infected voles resembled those of HCV infection in humans. In conclusion, infection with both BvHVs in their natural hosts shares striking similarities to HCV infection in humans and may represent promising small animal models for this important human disease.
Epigenetic targeting of SLC30A3 by HDAC1 is related to the malignant phenotype of glioblastoma

IUBMB life

2021 Mar 14

Zhang, L;Liu, Z;Dong, Y;Kong, L;
PMID: 33715270 | DOI: 10.1002/iub.2463

The epigenetic abnormality is believed as a major driver for cancer initiation. Histone modification plays a vital role in tumor formation and progression. Particularly, alteration in histone acetylation has been highly associated with gene expression, cell cycle, as well as carcinogenesis. By analyzing glioblastoma (GBM)-related microarray from the GEO database and conducting chromatin immunoprecipitation-sequencing (ChIP-seq), we discovered that solute carrier family 30 member 3 (SLC30A3), a super enhancer (SE)-regulated factor, was significantly reduced in GBM tissues. Furthermore, histone deacetylase 1 (HDAC1), overexpressed in GBM tissues, could inhibit SLC30A3 expression by promoting histone H3K27ac deacetylation modification of the SE region of SLC30A3. Our functional validation revealed that SLC30A3 can inhibit the growth and metastatic spread of GBM cells in vitro and in vivo, and can activate the MAPK signaling pathway to promote apoptosis of GBM cells. Moreover, overexpression of HDAC1 resulted in a significant increase in DNA replication activity, a significant decline in apoptosis and cell cycle arrest in GBM cells. In a word, these findings indicate that combined epigenetic targeting of SLC30A3 by HDAC1 and SE is potentially therapeutically feasible in GBM.
Ribonucleic Acid In Situ Hybridization Is a More Sensitive Method Than Immunohistochemistry in Detection of Thyroid Transcription Factor 1 and Napsin A Expression in Lung Adenocarcinomas.

Arch Pathol Lab Med.

2016 Apr 01

Shi J, Liu H, Ma XJ, Chen Z, He MX, Luo Y, Lin F1.
PMID: 27028392 | DOI: 10.5858/arpa.2014-0644-OA.

Abstract

CONTEXT:

-TTF-1 and napsin A immunomarkers have a crucial role in differentiating lung adenocarcinoma from lung squamous cell carcinoma and in identifying a primary lung adenocarcinoma when working on a tumor of unknown origin.

OBJECTIVES:

-To investigate the diagnostic sensitivity of ribonucleic acid in situ hybridization (RNAscope) in the detection of expression of these biomarkers in lung adenocarcinomas and to compare RNAscope to immunohistochemical techniques.

DESIGN:

-Both RNAscope and the immunohistochemical assays for TTF-1 and napsin A were performed on tissue microarray sections containing 80 lung adenocarcinomas and 80 lung squamous cell carcinomas. The RNAscope assay for both TTF-1 and napsin A was also performed on 220 adenocarcinomas from various organs.

RESULTS:

-The RNAscope assay for TTF-1 gave positive results in 92.5% (74 of 80) of the lung adenocarcinomas; in contrast, immunohistochemistry gave positive results in 82.5% (66 of 80) of those cases. The RNAscope assay for napsin A gave positive results in 90% (72 of 80) of lung adenocarcinomas; immunohistochemistry results were positive in 77.5% (62 of 80) of those cases. Napsin A expression was not seen in lung squamous cell carcinomas by either method. In contrast, TTF-1 expression was seen in 3.8% (3 of 80) (1(+)) and 10% (8 of 80) (1(+)) of the squamous cell carcinomas by immunochemistry and the RNAscope, respectively. All nonpulmonary adenocarcinoma results were negative for TTF-1 by the RNAscope assay.

CONCLUSIONS:

-Preliminary data suggest that RNAscope is superior to immunohistochemistry in detecting TTF-1 and napsin A expression in primary lung adenocarcinomas. Therefore, performing an RNAscope assay may be considered for both TTF-1(-) and napsin A(-) cases with a clinical suspicion of lung adenocarcinoma. The TTF-1 results should be interpreted with caution because a small percentage of squamous cell carcinomas can be focally positive by either assay.

In vitro and in vivo differences in neurovirulence between D614G, Delta And Omicron BA.1 SARS-CoV-2 variants

Acta neuropathologica communications

2022 Sep 05

Bauer, L;Rissmann, M;Benavides, FFW;Leijten, L;van Run, P;Begeman, L;Veldhuis Kroeze, EJB;Lendemeijer, B;Smeenk, H;de Vrij, FMS;Kushner, SA;Koopmans, MPG;Rockx, B;van Riel, D;
PMID: 36058935 | DOI: 10.1186/s40478-022-01426-4

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is associated with various neurological complications. Although the mechanism is not fully understood, several studies have shown that neuroinflammation occurs in the acute and post-acute phase. As these studies have predominantly been performed with isolates from 2020, it is unknown if there are differences among SARS-CoV-2 variants in their ability to cause neuroinflammation. Here, we compared the neuroinvasiveness, neurotropism and neurovirulence of the SARS-CoV-2 ancestral strain D614G, the Delta (B.1.617.2) and Omicron BA.1 (B.1.1.529) variants using in vitro and in vivo models. The Omicron BA.1 variant showed reduced neurotropism and neurovirulence compared to Delta and D614G in human induced pluripotent stem cell (hiPSC)-derived cortical neurons co-cultured with astrocytes. Similar differences were obtained in Syrian hamsters inoculated with D614G, Delta and the Omicron BA.1 variant 5 days post infection. Replication in the olfactory mucosa was observed in all hamsters, but most prominently in D614G inoculated hamsters. Furthermore, neuroinvasion into the CNS via the olfactory nerve was observed in D614G, but not Delta or Omicron BA.1 inoculated hamsters. Furthermore, neuroinvasion was associated with neuroinflammation in the olfactory bulb of hamsters inoculated with D614G. Altogether, our findings suggest differences in the neuroinvasive, neurotropic and neurovirulent potential between SARS-CoV-2 variants using in vitro hiPSC-derived neural cultures and in vivo in hamsters during the acute phase of the infection.
Cannabinoid CB2 receptors are expressed in glutamate neurons in the red nucleus and functionally modulate motor behavior in mice

Neuropharmacology

2021 Mar 28

Zhang, HY;Shen, H;Gao, M;Ma, Z;Hempel, B;Bi, GH;Gardner, EL;Wu, J;Xi, ZX;
PMID: 33789118 | DOI: 10.1016/j.neuropharm.2021.108538

Cannabinoids produce a number of central nervous system effects via the CB2 receptor (CB2R), including analgesia, antianxiety, anti-reward, hypoactivity and attenuation of opioid-induced respiratory depression. However, the cellular distributions of the CB2Rs in the brain remain unclear. We have reported that CB2Rs are expressed in midbrain dopamine (DA) neurons and functionally regulate DA-mediated behavior(s). Unexpectedly, high densities of CB2-like signaling were also found in a neighboring motor structure - the red nucleus (RN) of the midbrain. In the present study, we systematically explored CB2R expression and function in the RN. Immunohistochemistry and in situ hybridization assays showed high densities of CB2R-immunostaining and mRNA signal in RN magnocellular glutamate neurons in wildtype and CB1-knockout, but not CB2-knockout, mice. Ex vivo electrophysiological recordings in midbrain slices demonstrated that CB2R activation by JWH133 dose-dependently inhibited firing rates of RN magnocellular neurons in wildtype, but not CB2-knockout, mice, while having no effect on RN GABA neurons in transgenic GAD67-GFP reporter mice, suggesting CB2-mediated effects on glutamatergic neurons. In addition, microinjection of JWH133 into the RN produced robust ipsilateral rotations in wildtype, but not CB2-knockout mice, which was blocked by pretreatment with either a CB2 or DA D1 or D2 receptor antagonist, suggesting a DA-dependent effect. Finally, fluorescent tract tracing revealed glutamatergic projections from the RN to multiple brain areas including the ventral tegmental area, nucleus accumbens, thalamus, and cerebellum. These findings suggest that CB2Rs in RN glutamate neurons functionally modulate motor activity, and therefore, constitute a new target in cannabis-based medication development for motor disorders.
A role for paracrine interleukin-6 signaling in the tumor microenvironment in prostate tumor growth.

Prostate.

2018 Oct 21

Yu SH, Maynard JP, Vaghasia AM, De Marzo AM, Drake CG, Sfanos KS.
PMID: 30345534 | DOI: 10.1002/pros.23726

Abstract BACKGROUND: Interleukin-6 (IL-6) is a mediator of inflammation that can facilitate prostate cancer progression. We previously demonstrated that IL-6 is present in the prostate tumor microenvironment and is restricted almost exclusively to the stromal compartment. The present study examined the influence of paracrine IL-6 signaling on prostate tumor growth using allograft models of mouse prostate cancer (TRAMP-C2), colon cancer (MC38), and melanoma (B16) cell lines in wildtype (WT) and IL-6 knockout (IL-6-/- ) mice. METHODS: Cells were implanted into WT or IL-6-/- mice and tumor sizes were measured at a 3 to 4 day interval. Serum, tumors, and other organs were collected for IL-6 analysis by ELISA and RNA in situ hybridization (RISH). RESULTS: There was a significant reduction in TRAMP-C2 and B16 tumor size grown in IL-6-/- mice versus WT mice (P = 0.0006 and P = 0.02, respectively). This trend was not observed for the MC38 cell line. RISH analysis of TRAMP-C2 tumors grown in WT mice showed that cells present in the tumor microenvironment were the primary source of IL-6 mRNA, not the TRAMP-C2 cells. Serum IL-6 ELISA analyses showed an increase in the circulating levels of IL-6 in WT mice bearing TRAMP-C2 tumors. Similar phospho-STAT3 expression and tumor vascularization were observed in TRAMP-C2 tumors grown in WT and IL-6-/- mice. CONCLUSIONS: Our results are consistent with previous studies in prostate cancer patients demonstrating that paracrine IL-6 production in the tumor microenvironment may influence tumor growth. Additionally, these data provide evidence that elevated systemic IL-6 levels may be involved in tumor growth regulation in prostate cancer, and are not simply caused by or indicative of tumor burden.

EphA4 loss improves social memory performance and alters dendritic spine morphology without changes in amyloid pathology in a mouse model of Alzheimer's disease

Alzheimers Res Ther.

2019 Dec 12

Poppe L, Ru� L, Timmers M, Lenaerts A, Storm A, Callaerts-Vegh Z, Courtand G, de Boer A, Smolders S, Van Damme P, Van Den Bosch L, D'Hooge R, De Strooper B, Robberecht W, Lemmens R
PMID: 31831046 | DOI: 10.1186/s13195-019-0554-4

BACKGROUND: EphA4 is a receptor of the ephrin system regulating spine morphology and plasticity in the brain. These processes are pivotal in the pathophysiology of Alzheimer's disease (AD), characterized by synapse dysfunction and loss, and the progressive loss of memory and other cognitive functions. Reduced EphA4 signaling has been shown to rescue beta-amyloid-induced dendritic spine loss and long-term potentiation (LTP) deficits in cultured hippocampal slices and primary hippocampal cultures. In this study, we investigated whether EphA4 ablation might preserve synapse function and ameliorate cognitive performance in the APPPS1 transgenic mouse model of AD. METHODS: A postnatal genetic ablation of EphA4 in the forebrain was established in the APPPS1 mouse model of AD, followed by a battery of cognitive tests at 9 months of age to investigate cognitive function upon EphA4 loss. A Golgi-Cox staining was used to explore alterations in dendritic spine density and morphology in the CA1 region of the hippocampus. RESULTS: Upon EphA4 loss in APPPS1 mice, we observed improved social memory in the preference for social novelty test without affecting other cognitive functions. Dendritic spine analysis revealed altered synapse morphology as characterized by increased dendritic spine length and head width. These modifications were independent of hippocampal plaque load and beta-amyloid peptide levels since these were similar in mice with normal versus reduced levels of EphA4. CONCLUSION: Loss of EphA4 improved social memory in a mouse model of Alzheimer's disease in association with alterations in spine morphology.
Cutaneous lesions in psoriatic arthritis are enriched in chemokine transcriptomic pathways

Arthritis research & therapy

2023 May 02

Johnsson, H;Cole, J;Siebert, S;McInnes, IB;Graham, G;
PMID: 37131254 | DOI: 10.1186/s13075-023-03034-6

Skin from people with psoriasis has been extensively studied and is assumed to be identical to skin from those with psoriatic arthritis (PsA). Chemokines and the CC chemokine scavenger receptor ACKR2 are upregulated in uninvolved psoriasis. ACKR2 has been proposed as a regulator of cutaneous inflammation in psoriasis. The aim of this study was to compare the transcriptome of PsA skin to healthy control (HC) skin and evaluate ACKR2 expression in PsA skin.Full-thickness skin biopsies from HC, lesional and uninvolved skin from participants with PsA were sequenced on NovaSeq 6000. Findings were validated using qPCR and RNAscope.Nine HC and nine paired PsA skin samples were sequenced. PsA uninvolved skin was transcriptionally similar to HC skin, and lesional PsA skin was enriched in epidermal and inflammatory genes. Lesional PsA skin was enriched in chemokine-mediated signalling pathways, but uninvolved skin was not. ACKR2 was upregulated in lesional PsA skin but had unchanged expression in uninvolved compared with HC skin. The expression of ACKR2 was confirmed by qPCR, and RNAscope demonstrated strong expression of ACKR2 in the suprabasal layer of the epidermis in PsA lesions.Chemokines and their receptors are upregulated in lesional PsA skin but relatively unchanged in uninvolved PsA skin. In contrast to previous psoriasis studies, ACKR2 was not upregulated in uninvolved PsA skin. Further understanding of the chemokine system in PsA may help to explain why inflammation spreads from the skin to the joints in some people with psoriasis.
Changes in Hypothalamic Mu-Opioid Receptor Expression Following Acute Olanzapine Treatment in Female Rats: Implications for Feeding Behavior

Available at SSRN 

2023 May 15

Krogsbaek, M;Larsen, N;Landau, A;Sanchez, C;Nyengaard, J;
| DOI: 10.2139/ssrn.4436973

Advances have been made in recent years in using opioid receptor antagonists as an adjunct therapy to psychotropic medication to reduce debilitating weight gain and metabolic adverse effects associated with in particular second generation antipsychotics. However, it is unknown whether second generation antipsychotics produce a change in opioid receptor expression in the brain. The present study investigated early changes in opioid receptor expression in the female rat hypothalamus, a master controller of hunger and metabolic regulation, after acute treatment with olanzapine, a commonly used second generation antipsychotic. Using quantitative spatial in situ hybridization and receptor autoradiography, expression levels of the three opioid receptors; kappa, mu and delta, were determined at mRNA and protein level, respectively, in the five hypothalamic areas: paraventricular nucleus, arcuate nucleus, ventromedial nucleus, dorsomedial nucleus and lateral hypothalamus. After 48 hours of olanzapine treatment at clinically relevant plasma concentration weight gain and food intake changes, and increased plasma glucose were observed in female rats. Olanzapine treatment also led to a significant increase in mu opioid receptor availability in the arcuate nucleus, which contains both satiety and hunger controlling neurons. No other areas showed any opioid receptor expressional changes with olanzapine treatment on neither at mRNA nor protein level. Technical difficulties made it impossible to analyze mRNA levels in the lateral hypothalamus and overall binding of delta opioid receptors. Thus, the present study provided insights in to how olanzapine at clinically relevant plasma levels already at an early stage modulated the opioid system in the hypothalamus.
PP 2.15- 00169 Macrophages are the primary source of virus in semen and male genital tract organs in acutely and chronically infected rhesus macaques

Journal of Virus Eradication

2022 Dec 01

Deleage, C;Fennessey, C;Harper, J;Florea, S;Lipkey, L;Fast, R;Paiardini, M;Lifson, J;Keele, B;
| DOI: 10.1016/j.jve.2022.100170

Background: Most new HIV infections result from sexual interactions with infected but untreated individuals. Semen is the main vector for viral transmission globally, however, little is known regarding the anatomic origin and form of virus in semen. Methods: In this study, we were able to combine numerous new technologies to characterize the virus present in the semen during SIV infection. Six rhesus macaques (RM) were challenged intravenously with barcoded virus SIVmac239M. Semen and blood samples were collected longitudinally for 17 days post-infection with all male genital tract (MGT) and multiple lymphoid tissues collected at necropsy and subjected to quantitative PCR, next generation sequencing of the viral barcode, and tissue analysis (RNAscope, DNAscope and immunophenotyping). Semen was also collected from 6 animals chronically infected with SIVmac251 and in five CD4 depleted animals in acute phase and 2 weeks post ART initiation. Results: Extremely high levels of viral RNA (vRNA) were detected in seminal plasma (up to 10^9cp/ml) as well as comparable levels of cell associated vRNA and vDNA in seminal cells with detection starting as early as 4 days post-infection. RNAscope and immunophenotyping of seminal cells and MGT tissues revealed myeloid cells as the main source of virus (Fig. 1), while CD4+T cells were harboring vRNA in lymphoid tissues. Sequences show evidence of an early compartment between seminal and blood plasma and no difference in the env gene of virus present in semen/MGT and in Lymph Nodes. Finally, multinuclear giant cells harboring vRNA were the only source of virus in semen in chronically infected and in CD4 depleted RM. Moreover, vRNA + myeloid cells were highly present in semen after 2 weeks on ART.

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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
EnEmProbe targets exons n and m
En-EmProbe 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

Enabling research, drug development (CDx) and diagnostics

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