Polony gels enable amplifiable DNA stamping and spatial transcriptomics of chronic pain

Cell

2022 Nov 23

Fu, X;Sun, L;Dong, R;Chen, JY;Silakit, R;Condon, LF;Lin, Y;Lin, S;Palmiter, RD;Gu, L;
PMID: 36368323 | DOI: 10.1016/j.cell.2022.10.021

Methods for acquiring spatially resolved omics data from complex tissues use barcoded DNA arrays of low- to sub-micrometer features to achieve single-cell resolution. However, fabricating such arrays (randomly assembled beads, DNA nanoballs, or clusters) requires sequencing barcodes in each array, limiting cost-effectiveness and throughput. Here, we describe a vastly scalable stamping method to fabricate polony gels, arrays of ∼1-micrometer clonal DNA clusters bearing unique barcodes. By enabling repeatable enzymatic replication of barcode-patterned gels, this method, compared with the sequencing-dependent array fabrication, reduced cost by at least 35-fold and time to approximately 7 h. The gel stamping was implemented with a simple robotic arm and off-the-shelf reagents. We leveraged the resolution and RNA capture efficiency of polony gels to develop Pixel-seq, a single-cell spatial transcriptomic assay, and applied it to map the mouse parabrachial nucleus and analyze changes in neuropathic pain-regulated transcriptomes and cell-cell communication after nerve ligation.

Recalibrating vision-for-action requires years after sight restoration from congenital cataracts

eLife

2022 Oct 24

Senna, I;Piller, S;Ben-Zion, I;Ernst, MO;
PMID: 36278872 | DOI: 10.7554/eLife.78734

Being able to perform adept goal-directed actions requires predictive, feed-forward control, including a mapping between the visually estimated target locations and the motor commands reaching for them. When the mapping is perturbed, e.g., due to muscle fatigue or optical distortions, we are quickly able to recalibrate the sensorimotor system to update this mapping. Here, we investigated whether early visual and visuomotor experience is essential for developing sensorimotor recalibration. To this end, we assessed young individuals deprived of pattern vision due to dense congenital bilateral cataracts who were surgically treated for sight restoration only years after birth. We compared their recalibration performance to such distortion to that of age-matched sighted controls. Their sensorimotor recalibration performance was impaired right after surgery. This finding cannot be explained by their still lower visual acuity alone, since blurring vision in controls to a matching degree did not lead to comparable behavior. Nevertheless, the recalibration ability of cataract-treated participants gradually improved with time after surgery. Thus, the lack of early pattern vision affects visuomotor recalibration. However, this ability is not lost but slowly develops after sight restoration, highlighting the importance of sensorimotor experience gained late in life.

Orexin neurons inhibit sleep to promote arousal

Nature communications

2022 Jul 18

De Luca, R;Nardone, S;Grace, KP;Venner, A;Cristofolini, M;Bandaru, SS;Sohn, LT;Kong, D;Mochizuki, T;Viberti, B;Zhu, L;Zito, A;Scammell, TE;Saper, CB;Lowell, BB;Fuller, PM;Arrigoni, E;
PMID: 35851580 | DOI: 10.1038/s41467-022-31591-y

Humans and animals lacking orexin neurons exhibit daytime sleepiness, sleep attacks, and state instability. While the circuit basis by which orexin neurons contribute to consolidated wakefulness remains unclear, existing models posit that orexin neurons provide their wake-stabilizing influence by exerting excitatory tone on other brain arousal nodes. Here we show using in vivo optogenetics, in vitro optogenetic-based circuit mapping, and single-cell transcriptomics that orexin neurons also contribute to arousal maintenance through indirect inhibition of sleep-promoting neurons of the ventrolateral preoptic nucleus. Activation of this subcortical circuit rapidly drives wakefulness from sleep by differentially modulating the activity of ventrolateral preoptic neurons. We further identify and characterize a feedforward circuit through which orexin (and co-released glutamate) acts to indirectly target and inhibit sleep-promoting ventrolateral preoptic neurons to produce arousal. This revealed circuitry provides an alternate framework for understanding how orexin neurons contribute to the maintenance of consolidated wakefulness and stabilize behavioral state.

Development of Glioblastoma from Stem Cells to a Full-Fledged Tumor

Turk patoloji dergisi

2022 Jun 27

Vladimirovich, NP;Railevna, MG;Nikolaevich, PV;Nikolaevich, GD;Mikhailovich, KV;Leonard, W;Mauric, K;Sergeevich, TP;
PMID: 35876685 | DOI: 10.5146/tjpath.2022.01582

IDH wild-type glioblastomas (GBM) are one of the most malignant and complex tumors for treatment. The urgent question of new therapeutic and diagnostic tools searching should be resolved based on cellular and molecular pathogenesis mechanisms, which remain insufficiently studied. In this study, we aimed to investigate GBM pathogenesis.Using the isolation of different GBM cell populations and the cell cultures, animal models, and molecular genetic methods, we tried to clarify the picture of GBM pathogenesis by constructing a projection from different glioma stem cells types to an integral neoplasm.We have shown a potential transformation pathway for both glioma stem cells and four definitive cell populations during gliomagenesis. Moreover, we have characterized each population, taking into account its place in the pathogenetic continuum, with a description of the most fundamental molecular and functional properties.Finally, we have formed a complex holistic concept of the pathogenetic evolution of GBM at the cell-population level by integrating our results with the data of the world literature.

Inhibition of ATP-citrate lyase improves NASH, liver fibrosis, and dyslipidemia

Cell metabolism

2022 Jun 07

Morrow, MR;Batchuluun, B;Wu, J;Ahmadi, E;Leroux, JM;Mohammadi-Shemirani, P;Desjardins, EM;Wang, Z;Tsakiridis, EE;Lavoie, DCT;Reihani, A;Smith, BK;Kwiecien, JM;Lally, JSV;Nero, TL;Parker, MW;Ask, K;Scott, JW;Jiang, L;Paré, G;Pinkosky, SL;Steinberg, GR;
PMID: 35675800 | DOI: 10.1016/j.cmet.2022.05.004

Elevated liver de novo lipogenesis contributes to non-alcoholic steatohepatitis (NASH) and can be inhibited by targeting acetyl-CoA carboxylase (ACC). However, hypertriglyceridemia limits the use of pharmacological ACC inhibitors as a monotherapy. ATP-citrate lyase (ACLY) generates acetyl-CoA and oxaloacetate from citrate, but whether inhibition is effective for treating NASH is unknown. Here, we characterize a new mouse model that replicates many of the pathological and molecular drivers of NASH and find that genetically inhibiting ACLY in hepatocytes reduces liver malonyl-CoA, oxaloacetate, steatosis, and ballooning as well as blood glucose, triglycerides, and cholesterol. Pharmacological inhibition of ACLY mirrors genetic inhibition but has additional positive effects on hepatic stellate cells, liver inflammation, and fibrosis. Mendelian randomization of human variants that mimic reductions in ACLY also associate with lower circulating triglycerides and biomarkers of NASH. These data indicate that inhibiting liver ACLY may be an effective approach for treatment of NASH and dyslipidemia.

Cellular Architecture of Human Brain Metastases

SSRN Electronic Journal

2021 Jul 22

Gonzalez, H;Mei, W;Robles, I;Hagerling, C;Allen, B;Nanjaraj, A;Verbeek, T;Kalavacherla, S;van Gogh, M;Georgiou, S;Daras, M;Philips, J;Spitzer, M;Roose, J;Werb, Z;
| DOI: 10.2139/ssrn.3883639

Brain metastasis (BrM) is the most common form of brain cancer, characterized by neurologic disability and abysmal prognosis. Unfortunately, our understanding of the biology underlying human BrMs remains rudimentary. Here we present an integrative analysis of >100,000 malignant and non-malignant cells from 15 human parenchymal BrMs as a resource, generated by single-cell transcriptomics, mass cytometry, and complemented with mouse model- and in silico- approaches. We interrogated the composition of BrM niches, molecularly defined the blood-tumor interface, and reveal stromal immunosuppressive states, enriched with infiltrated T cells and macrophages. Specific single-cell interrogation of metastatic tumor cells provides a novel framework of 8 functional cell programs that coexist or anticorrelate. Collectively, these programs delineate two functional BrM archetypes, one proliferative and the other inflammatory, that are evidently shaped through tumor-immune interactions. Our resource provides a foundation to understand the molecular basis of BrM in patients with tumor cell-intrinsic and host environmental- traits.

Clinicopathological and molecular characterization of high-grade endometrial carcinoma with POLE mutation: a single center study

Journal of Gynecologic Oncology

2022 Feb 08

Yu, S;Sun, Z;Zong, L;Yan, J;Yu, M;Chen, J;Lu, Z;
| DOI: 10.3802/jgo.2022.33.e38

Patients with POLEmut generally have early-stage disease [5]; in our study, 93% (40/43) of all POLEmut patients had stage I or II EC, which was consistent with previously reported data [19]. In terms of treatment strategy, our data suggest that the type of adjuvant therapy after surgery does not impact the survival of patients with POLEmut ECs who are FIGO stage I-II disease, which is consistent with the ESGO/ESTRO/ESP clinical practice guidelines [15]; as such, these data maybe support omitting additional adjuvant therapy for such patients, besides, the recent meta-analysis from a 294-patients group also give the similar conclusion [20]. However, FIGO stage III and IV patients with POLEmut are rare [5]; our cohort comprised only 1 patient each with stage III and stage IV (Fig. 1B). The individual with stage III had a very good prognosis (i.e., no recurrence or death 40 months after surgery), but the individual with stage IV had a poor prognosis (Fig. 2, Tables S1 and S2, patient #3). The 2021 ESGO/ESTRO/ESP guidelines define patients with stage III-IV disease with residual tumor as advanced risk, regardless of molecular type [15]. However, no recommendations or instructions are provided for patients with stage III-IVA disease exhibiting POLEmut without residual tumors because of limited data.

Crosstalk Between Pro-Survival Sphingolipid Metabolism and Complement Signaling Induces Inflammasome-Mediated Tumor Metastasis

SSRN Electronic Journal

2022 Feb 17

Janneh, A;Kassir, M;Atilgan, F;Lee, H;Sheridan, M;Oleinik, N;Szulc, Z;Voelkel-Johnson, C;Nguyen, H;Li, H;Peterson, Y;Marangoni, E;Saatci, O;Sahin, O;Lilly, M;Atkinson, C;Tomlinson, S;Mehrotra, S;Ogretmen, B;
| DOI: 10.2139/ssrn.4032074

Crosstalk between metabolic and signaling events that induce tumor metastasis remains elusive. Here, we determined how oncogenic sphingosine 1-phosphate (S1P) metabolism activates intracellular C3-complement molecules to enhance migration/metastasis. We demonstrate that increased S1P metabolism activates C3-complement processing through S1P receptor 1 (S1PR1). S1P/S1PR1-activated intracellular C3b-α′2 was associated with PPIL1 through glutamic acid 156 (E156) and aspartic acid 111 (D111) residues, resulting in NLRP3/inflammasome induction. Inactivation mutations of S1PR1 to prevent S1P signaling or mutations of C3b-α′2 to prevent its association with PPIL1 attenuated inflammasome activation and reduced lung colonization/metastasis in mice. Also, activation of the S1PR1/C3/PPIL1/NLRP3 axis was highly associated with human metastatic melanoma tissues and patient-derived xenografts (PDXs). Moreover, targeting S1PR1/C3/PPIL1/NLRP3 signaling using molecular, genetic, and/or pharmacologic tools prevented lung colonization/metastasis of B16 melanoma-derived tumors using WT and C3a-receptor1 knockout (C3aR1-/-) mice. Overall, these data provide novel strategies for the treatment of high-grade/metastatic tumors by targeting the S1PR1/C3/inflammasome axis.

Social touch-like tactile stimulation activates a tachykinin 1-oxytocin pathway to promote social interactions

Neuron

2022 Jan 12

Yu, H;Miao, W;Ji, E;Huang, S;Jin, S;Zhu, X;Liu, MZ;Sun, YG;Xu, F;Yu, X;
PMID: 35045339 | DOI: 10.1016/j.neuron.2021.12.022

It is well known that affective and pleasant touch promotes individual well-being and facilitates affiliative social communication, although the neural circuit that mediates this process is largely unknown. Here, we show that social-touch-like tactile stimulation (ST) enhances firing of oxytocin neurons in the mouse paraventricular hypothalamus (PVH) and promotes social interactions and positively reinforcing place preference. These results link pleasant somatosensory stimulation to increased social interactions and positive affective valence. We further show that tachykinin 1 (Tac1+) neurons in the lateral and ventrolateral periaqueductal gray (l/vlPAG) send monosynaptic excitatory projections to PVH oxytocin neurons. Functionally, activation of PVH-projecting Tac1+ neurons increases firing of oxytocin neurons, promotes social interactions, and increases preference for the social touch context, whereas reducing activity of Tac1+ neurons abolishes ST-induced oxytocin neuronal firing. Together, these results identify a dipeptidergic pathway from l/vlPAG Tac1+ neurons to PVH oxytocin neurons, through which pleasant sensory experience promotes social behavior.

Acute restraint stress redirects prefrontal cortex circuit function through mGlu5 receptor plasticity on somatostatin-expressing interneurons

Neuron

2022 Jan 13

Joffe, ME;Maksymetz, J;Luschinger, JR;Dogra, S;Ferranti, AS;Luessen, DJ;Gallinger, IM;Xiang, Z;Branthwaite, H;Melugin, PR;Williford, KM;Centanni, SW;Shields, BC;Lindsley, CW;Calipari, ES;Siciliano, CA;Niswender, CM;Tadross, MR;Winder, DG;Conn, PJ;
PMID: 35045338 | DOI: 10.1016/j.neuron.2021.12.027

Inhibitory interneurons orchestrate prefrontal cortex (PFC) activity, but we have a limited understanding of the molecular and experience-dependent mechanisms that regulate synaptic plasticity across PFC microcircuits. We discovered that mGlu5 receptor activation facilitates long-term potentiation at synapses from the basolateral amygdala (BLA) onto somatostatin-expressing interneurons (SST-INs) in mice. This plasticity appeared to be recruited during acute restraint stress, which induced intracellular calcium mobilization within SST-INs and rapidly potentiated postsynaptic strength onto SST-INs. Restraint stress and mGlu5 receptor activation each augmented BLA recruitment of SST-IN phasic feedforward inhibition, shunting information from other excitatory inputs, including the mediodorsal thalamus. Finally, studies using cell-type-specific mGlu5 receptor knockout mice revealed that mGlu5 receptor function in SST-expressing cells is necessary for restraint stress-induced changes to PFC physiology and related behaviors. These findings provide new insights into interneuron-specific synaptic plasticity mechanisms and suggest that SST-IN microcircuits may be promising targets for treating stress-induced psychiatric diseases.

Nasopharyngeal cancer: Incidence and prognosis of human papillomavirus and Epstein-Barr virus association at a single North American institution

Head & neck

2022 Jan 18

Wu, SS;Chen, B;Fleming, CW;Shah, AA;Griffith, CC;Domb, C;Reddy, CA;Campbell, SR;Woody, NM;Lamarre, ED;Lorenz, RR;Prendes, BL;Scharpf, J;Schwartzman, L;Geiger, JL;Koyfman, SA;Ku, JA;
PMID: 35040516 | DOI: 10.1002/hed.26976

The prognostication of Epstein-Barr virus (EBV) and human papillomavirus (HPV) status in nasopharyngeal cancer (NPC) is unclear.This retrospective study analyzed NPC from 2000 to 2019.Seventy-eight patients were included: 43 EBV+ , 12 HPV+ , 23 EBV- /HPV- , and 0 EBV+ /HPV+ . All p16+ tumors were also positive for HPV-CISH. Baseline characteristics were not different between groups except age, N-classification, and Karnofsky Performance Scale (KPS) (p < 0.05). For EBV+ , HPV+ , and EBV- /HPV- respectively, 3-year overall survival (OS) was 89.9%, 69.8%, and 52.5% (p = 0.006). EBV- /HPV- status was significantly associated with worse OS but not freedom from progression (FFP) on univariate analysis, and did not remain a significant predictor of OS after adjusting for KPS, age, and group stage.EBV+ NPC tumors were seen in younger, healthier patients than HPV+ and EBV- tumors, and there were no cases of coinfection. The association of viral status with OS was insignificant after adjusting for KPS and age.

The assembly of caprine Y chromosome sequence reveals a unique paternal phylogenetic pattern and improves our understanding of the origin of domestic goat

Ecology and evolution

2021 Jun 01

Xiao, C;Li, J;Xie, T;Chen, J;Zhang, S;Elaksher, SH;Jiang, F;Jiang, Y;Zhang, L;Zhang, W;Xiang, Y;Wu, Z;Zhao, S;Du, X;
PMID: 34188851 | DOI: 10.1002/ece3.7611

The mammalian Y chromosome offers a unique perspective on the male reproduction and paternal evolutionary histories. However, further understanding of the Y chromosome biology for most mammals is hindered by the lack of a Y chromosome assembly. This study presents an integrated in silico strategy for identifying and assembling the goat Y-linked scaffolds using existing data. A total of 11.5 Mb Y-linked sequences were clustered into 33 scaffolds, and 187 protein-coding genes were annotated. We also identified high abundance of repetitive elements. A 5.84 Mb subset was further ordered into an assembly with the evidence from the goat radiation hybrid map (RH map). The existing whole-genome resequencing data of 96 goats (worldwide distribution) were utilized to exploit the paternal relationships among bezoars and domestic goats. Goat paternal lineages were clearly divided into two clades (Y1 and Y2), predating the goat domestication. Demographic history analyses indicated that maternal lineages experienced a bottleneck effect around 2,000 YBP (years before present), after which goats belonging to the A haplogroup spread worldwide from the Near East. As opposed to this, paternal lineages experienced a population decline around the 10,000 YBP. The evidence from the Y chromosome suggests that male goats were not affected by the A haplogroup worldwide transmission, which implies sexually unbalanced contribution to the goat trade and population expansion in post-Neolithic period.

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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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