Journal: Cells

Article Title: Disruption of Circadian Rhythms by Ambient Light during Neurodevelopment Leads to Autistic-like Molecular and Behavioral Alterations in Adult Mice

doi: 10.3390/cells10123314

Figure Lengend Snippet: Daily rhythms of animal locomotor activities and clock gene expression are disrupted in SD mice. ( A ) A diagram indicating the strategy to generate the SD mice. ( B ) Top: Representative actograms of mouse wheel-running activities in control (CTR) and Short Day (SD) mice. Middle: Morlet wavelet transform analysis below indicates that CTR mice entrained to the 12 h/12 h light/dark cycle and exhibited a circadian period of 24 h. In contrast, SD mice entrained to the 8 h/8 h light/dark cycle and exhibited a circadian period of 8 h. Bottom: Average activities of CTR (n = 4) and SD (n = 3) mice across 24 h. Note that the SD mice exhibited a higher level of total activities in 24 h as compared with the CTR mice. ( C ) Representative microscopic images of immunostaining for PER1 and PER2 in the SCN. ZT1: one hour after light-on for CTR and SD mice; ZT13 and ZT9: 1 h after light-off for CTR and SD mice. Quantitative analysis of staining intensity is shown to the right. The levels at ZT13 and ZT9 were normalized according to the levels at ZT1 in the CTR and the SD mice, respectively. Note that PER1 and Per2 levels were not different between ZT1 and ZT9 in SD mice. Data are displayed as individual values and mean ± SEM. n = 3 in CTR and n = 4–5 in SD. **** p < 0.0001; n.s., not significant vs. ZT1. ( D ) Western blotting images indicate that PER1, PER2, and BMAL1 exhibited circadian oscillations in the hippocampi of the CTR, but not the SD, mice. Quantitation of protein levels is shown below. For this experiment mice were kept in constant darkness for 48 h and hippocampal samples were harvested every 4 h in the next 24 h. Data are displayed as mean ± SEM. n = 3 in CTR and SD. ** p < 0.01; * p < 0.05 vs. CTR.

Article Snippet: Immunohistochemistry was performed as published in [ ], using one of the following primary antibodies: rabbit anti-phospho-eIF4E (Ser209) antibody (1:1000 final dilution; Novus Biologicals, NBP1-19923; RRID: AB_1641951); rabbit anti-phospho-S6 (Ser240/244) antibody (1:1000 final dilution; Cell Signaling Tech, 2215); rabbit anti-PER1 antibody (1:3000, Millipore, AB2201); or goat anti-PER2 antibody (1:300, Santa Cruz, sc-7728).

Techniques: Gene Expression, Control, Immunostaining, Staining, Western Blot, Quantitation Assay

Journal: Cell cycle (Georgetown, Tex.)

Article Title: Effect of circadian clock mutations on DNA damage response in mammalian cells.

doi: 10.4161/cc.21771

Figure Lengend Snippet: Figure 5. Mouse cells deficient in clock genes are not sensitive to mitomycin C. Cells were plated at a low density, allowed to attach and then treated with mitomy- cin C (MMC); after colony formation, cells were stained with Giemsa, and percent survival was determined as described in the text. Each data point represents the average of at least three independent experiments and bars signify the standard deviation. (A) Mouse embryonic fibroblasts were derived from mice with the following genotypes: wild-type (squares), Clock-/- (circles) and Bmal1-/- (triangles). (B) Mouse skin fibroblasts were derived from mice with the following genotypes: wild-type (squares), Cry1-/-Cry2-/- (circles) and Per1-/-Per2-/- (triangles).

Article Snippet: For siRNA transfections, exponentially growing cells were transfected either with human Per1, Per2, mouse Per1, Per2, Cry1, Cry2, Clock and Bmal1, and Cyclophilin B siRNAs (Santa Cruz) or non-target siRNA (Dharmacon) using the Lipofectamine RNAiMax (Invitrogen) transfection reagent for 48 h according to the manufacturer’s directions before UV or IR irradiation. pCDNA3 or V5- tagged mouse Per1 genes were transfected using the Lipofectamine 2000 (Invitrogen) transfection reagent for 48 h according to the manufacturer’s directions as described previously.40 Clonogenic UV survival assays.

Techniques: Staining, Standard Deviation, Derivative Assay

Journal: Alzheimer's & dementia : the journal of the Alzheimer's Association

Article Title: Tauopathy after long-term cervical lymphadenectomy.

doi: 10.1002/alz.70136

Figure Lengend Snippet: FIGURE 2 Long-term cervical lymphadenectomy (cLE) leads to sleep disorders. A, The volcano plot illustrates the number of upregulated and downregulated differentially expressed genes in brain tissue bulk RNA sequencing results after cLE (p < 0.01, n = 6 mice per group). B, Intersection of all differentially expressed genes with depressive disorder-related genes identifying eight overlapping genes. C, Differential expression levels of the eight overlapping genes: Pomc, Dusp1, Npas2, Mmab, Map2k5, Atp2a2, Per3, and Per2. D, Kyoto Encyclopedia of Genes and Genomes (KEGG)

Article Snippet: Primary antibodies used include: Per2 Rabbit pAb (Abclonal, A13168, 1:1000 for Western Blotting); Npas2 Rabbit pAb (Abclonal, A16930, 1:1000 for western blotting); Pomc Rabbit pAb (Abclonal, A3038, 1:1000 for western blotting, 1:100 for immunofluorescence); NeuN mouse mAb (Millipore, MAB377, 1:200 for immunofluorescence); pMEK1/2 Rabbit mAb (Cell Signaling Technology, #2338, 1:2000 for western blotting); p-Erk1/2 Rabbit mAb (Cell Signaling Technology, #4370, 1:2000 for western blotting); t-Erk1/2 Rabbit pAb (Cell Signaling Technology, #9102, 1:1000 for western blotting); p-p38 Rabbit mAb (Cell Signaling Technology, #4511, 1:1000 for western blotting); LC3B Rabbit mAb (Cell Signaling Technology, #3868, 1:1000 for western blotting); Atg12 Rabbit mAb (Abclonal, A22788, 1:500 for western blotting); Beclin1 Rabbit mAb (Abclonal, A21191, 1:5000 for western blotting); p62 Rabbit mAb (Abclonal, A19700, 1:20000 for western blotting); Ki67 Rabbit mAb (Abcam, ab16667, 1:1000 for western blotting, 1:200 for immunofluorescence); APP Rabbit mAb (Abcam, ab32136, 1:10000 for western blotting); Aβ1-42 Rabbit mAb (Abcam, ab201060, 1:5000 for western blotting); p-tau205 Rabbit mAb (Abclonal, AP1445, 1:500 for western blotting, 1:100 15525279, 2025, 4, D ow nloaded from https://alz-journals.onlinelibrary.w iley.com /doi/10.1002/alz.70136, W iley O nline L ibrary on [10/04/2025].

Techniques: RNA Sequencing, Quantitative Proteomics

Journal: International Journal of Oral Science

Article Title: PER2-mediated ameloblast differentiation via PPARγ/AKT1/β-catenin axis

doi: 10.1038/s41368-021-00123-7

Figure Lengend Snippet: Dysregulated signalling molecules in the ameloblasts of neonatal offspring of circadian disruption pregnant mice. An environmental circadian disruption model was constructed in 10- to 12-week-old pregnant mice. a Total protein of mandibular first molar germs of the offspring (PN3) was extracted. Compared to the control group, the protein levels of PER2, PPARγ and AMELX decreased in the disturbance group. b The mandibles of the offspring (PN5) were dissected. Compared to the control group, the expression levels of PER2, PPARγ, AKT1-Ser473 and β-catenin-Ser552 were reduced in ameloblasts in the disturbance group. ** P < 0.01; *** P < 0.001. Bar = 20 μm

Article Snippet: Primary antibodies included PER2 (NBP2-24616, Novus Biologicals), BMAL1 (NB100-2288, Novus Biologicals), PPARγ (A0270, ABclonal Technology, China), AKT1-Ser473 (AP0140, ABclonal Technology, China), β-catenin-Ser552 (AP0579, ABclonal Technology, China), AKT1 (A11016, ABclonal Technology, China), AMELX (ab153915, Abcam), β-catenin (A11343, ABclonal Technology, China), LAMIN B1 (A1910, ABclonal Technology, China), β-tubulin (PMK081M, BioPM, China), GAPDH (PMK042M, BioPM, China), CK14 (MA5-11599, ThermoFisher Scientific), F-actin (ab205, Abcam), and E-cadherin (sc-8426, Santa Cruz).

Techniques: Disruption, Construct, Control, Expressing

Journal: International Journal of Oral Science

Article Title: PER2-mediated ameloblast differentiation via PPARγ/AKT1/β-catenin axis

doi: 10.1038/s41368-021-00123-7

Figure Lengend Snippet: Altered signalling molecules expression and differentiation inhibition in Per2 -knockdown ALC cells. a Construction of Per2 knockdown ALC cell line. The knockdown efficiency of ALC- Per2 -sh was examined by qRT-PCR and western blot. b , c Expression of PPARγ, AMELX, AKT1-Ser473 and β-catenin-Ser552 was reduced in ALC- Per2 -sh. d , e ALC-Con cells and ALC- Per2 -sh cells were cultured in differentiation-inducing medium. On days 3, 7, 14 and 21 of differentiation induction, in ALC- Per2 -sh cells, ALP staining weakened ( e ), and the transcription levels of Alp and Ocn decreased ( d ). * P < 0.05; ** P < 0.01; *** P < 0.001

Article Snippet: Primary antibodies included PER2 (NBP2-24616, Novus Biologicals), BMAL1 (NB100-2288, Novus Biologicals), PPARγ (A0270, ABclonal Technology, China), AKT1-Ser473 (AP0140, ABclonal Technology, China), β-catenin-Ser552 (AP0579, ABclonal Technology, China), AKT1 (A11016, ABclonal Technology, China), AMELX (ab153915, Abcam), β-catenin (A11343, ABclonal Technology, China), LAMIN B1 (A1910, ABclonal Technology, China), β-tubulin (PMK081M, BioPM, China), GAPDH (PMK042M, BioPM, China), CK14 (MA5-11599, ThermoFisher Scientific), F-actin (ab205, Abcam), and E-cadherin (sc-8426, Santa Cruz).

Techniques: Expressing, Inhibition, Knockdown, Quantitative RT-PCR, Western Blot, Cell Culture, Staining

Journal: International Journal of Oral Science

Article Title: PER2-mediated ameloblast differentiation via PPARγ/AKT1/β-catenin axis

doi: 10.1038/s41368-021-00123-7

Figure Lengend Snippet: β-catenin translocated into the nucleus, and the subcellular localization of E-cadherin changed in Per2 -knockdown ALC cells. a , b ALC-Con cells and ALC- Per2 -sh cells were cultured in differentiation-inducing medium. On days 7, 14 and 21 of differentiation induction, in ALC- Per2 -sh cells, β-catenin expression increased in the nucleus ( a ) and decreased in the cytoplasm ( b ) compared to that in control cells; c Per2 knockdown altered the subcellular localization of E-cadherin in ALC cells. Original magnification, ×100. * P < 0.05; ** P < 0.01

Article Snippet: Primary antibodies included PER2 (NBP2-24616, Novus Biologicals), BMAL1 (NB100-2288, Novus Biologicals), PPARγ (A0270, ABclonal Technology, China), AKT1-Ser473 (AP0140, ABclonal Technology, China), β-catenin-Ser552 (AP0579, ABclonal Technology, China), AKT1 (A11016, ABclonal Technology, China), AMELX (ab153915, Abcam), β-catenin (A11343, ABclonal Technology, China), LAMIN B1 (A1910, ABclonal Technology, China), β-tubulin (PMK081M, BioPM, China), GAPDH (PMK042M, BioPM, China), CK14 (MA5-11599, ThermoFisher Scientific), F-actin (ab205, Abcam), and E-cadherin (sc-8426, Santa Cruz).

Techniques: Knockdown, Cell Culture, Expressing, Control

Journal: International Journal of Oral Science

Article Title: PER2-mediated ameloblast differentiation via PPARγ/AKT1/β-catenin axis

doi: 10.1038/s41368-021-00123-7

Figure Lengend Snippet: Overexpression of PPARγ partially rescued the altered signalling molecule expression and weakened ALP staining and ALP activity in Per2 -knockdown ALC cells. a PPARγ was overexpressed in ALC- Per2 -sh cells transfected with plasmids. With the increase in PPARγ expression, PER2 expression remained unchanged, while AKT1 and β-catenin phosphorylation levels were enhanced in the ALC- Per2 -sh-pEnCMV- Pparγ group. b ALP staining and ALP activity were enhanced in ALC- Per2 -sh-pEnCMV- Pparγ cells compared to those in ALC- Per2 -sh-pEnCMV cells but still weakened compared to those in ALC-Con-pEnCMV cells. * P < 0.05; ** P < 0.01; *** P < 0.001

Article Snippet: Primary antibodies included PER2 (NBP2-24616, Novus Biologicals), BMAL1 (NB100-2288, Novus Biologicals), PPARγ (A0270, ABclonal Technology, China), AKT1-Ser473 (AP0140, ABclonal Technology, China), β-catenin-Ser552 (AP0579, ABclonal Technology, China), AKT1 (A11016, ABclonal Technology, China), AMELX (ab153915, Abcam), β-catenin (A11343, ABclonal Technology, China), LAMIN B1 (A1910, ABclonal Technology, China), β-tubulin (PMK081M, BioPM, China), GAPDH (PMK042M, BioPM, China), CK14 (MA5-11599, ThermoFisher Scientific), F-actin (ab205, Abcam), and E-cadherin (sc-8426, Santa Cruz).

Techniques: Over Expression, Expressing, Staining, Activity Assay, Knockdown, Transfection, Phospho-proteomics

Journal: International Journal of Oral Science

Article Title: PER2-mediated ameloblast differentiation via PPARγ/AKT1/β-catenin axis

doi: 10.1038/s41368-021-00123-7

Figure Lengend Snippet: Overexpression of PPARγ reversed β-catenin subcellular localization in Per2 -knockdown ALC cells. ALC-Con and ALC- Per2 -sh were transfected with plasmids. a , b In the differentiation assay, the expression of β-catenin was obviously decreased in the nucleus ( a ) and cytoplasm ( b ) in ALC- Per2 -sh-pEnCMV- Pparγ cells. c Cell immunofluorescence showed that β-catenin translocated into the nucleus in ALC- Per2 -sh-pEnCMV cells compared with ALC-Con-pEnCMV cells but was reversed in ALC- Per2 -sh-pEnCMV- Pparγ cells. * P < 0.05; ** P < 0.01; *** P < 0.001

Article Snippet: Primary antibodies included PER2 (NBP2-24616, Novus Biologicals), BMAL1 (NB100-2288, Novus Biologicals), PPARγ (A0270, ABclonal Technology, China), AKT1-Ser473 (AP0140, ABclonal Technology, China), β-catenin-Ser552 (AP0579, ABclonal Technology, China), AKT1 (A11016, ABclonal Technology, China), AMELX (ab153915, Abcam), β-catenin (A11343, ABclonal Technology, China), LAMIN B1 (A1910, ABclonal Technology, China), β-tubulin (PMK081M, BioPM, China), GAPDH (PMK042M, BioPM, China), CK14 (MA5-11599, ThermoFisher Scientific), F-actin (ab205, Abcam), and E-cadherin (sc-8426, Santa Cruz).

Techniques: Over Expression, Knockdown, Transfection, Differentiation Assay, Expressing, Immunofluorescence

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: Integrative bioinformatics and experimental validation identify circadian clock component PER2 as a key regulator in asthma. A Venn diagram of significant genes from GSE63142 and CRGs. B The Random Forest algorithm was selected 11 genes based on Mean Decrease Accuracy scores greater than 2.0. C The LASSO logistic regression algorithm determined 16 genes. D The PPI network analysis identified top 10 genes by the CytoHubba plugin. E Venn diagram of the overlapping hub genes between the Random Forest, LASSO algorithm and PPI network. F - G Association of PER2 and PER3 mRNA expression levels in bronchial epithelial cells between control ( n = 27), mild-moderate asthma (MMA) ( n = 72), and severe asthma (SA) ( n = 56) patients from GSE63142 . PER2 and PER3 mRNA expression levels have been logarithm base 2 (log2)-transformed. H - I Association of PER2 and PER3 mRNA expression levels in bronchial epithelial brushings between control ( n = 30), and patients with asthma ( n = 51) from GSE41861 . PER2 and PER3 mRNA expression levels have been log2-transformed. J - K Association of PER2 and PER3 mRNA expression levels in bronchial epithelial cells between control ( n = 20), mild-moderate asthma (MMA) ( n = 50), and severe asthma (SA) ( n = 38) patients from GSE43696 . PER2 and PER3 mRNA expression levels have been log2-transformed. L - O Correlation between PER2 mRNA expression in primary bronchial epithelial cells and FEV 1 (L), FEV 1 % predicted, FVC (L), IgE (IU/mL) was determined by Spearman analysis in patients with asthma from GSE201955 ( n = 79). P qPCR analysis of the expression of Per2 in lung tissues of mice ( n = 3). Q - R Western blot analysis the expression of PER2 in lung tissues of mice treated with OVA or PBS ( n = 3). S - T Representative Immunofluorescence staining of PER2 (Red) in the airway epithelium (SCGB1A1 Green) from asthmatic and control mice ( n = 3), original magnification 200 ×. Scale bars, 100 μm. Statistical significance was determined using the Kruskal-Wallis test followed by turkey’s comparison test for ( F , G , J , K ); Mann-Whitney U test for ( H , I ); spearman correlation for ( L - O ); and unpaired Student’s t-test for ( P , R , T ). CRG, circadian related genes; LASSO, least absolute shrinkage and selection operator; RF, random forest; PPI, protein-protein interaction; MMA, mild-moderate asthma; SA, severe asthma; FEV 1 , forced expiratory volume in 1 s; FVC, forced vital capacity; IgE, immunoglobulin E. ** P < 0.01, *** P < 0.001

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: Biomarker Discovery, Expressing, Control, Transformation Assay, Western Blot, Immunofluorescence, Staining, Comparison, MANN-WHITNEY, Selection

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: Per2 −/− mice aggravates OVA-induced AHR, airway inflammation, mucus production and fibrosis in experimental asthma. A Schematic illustrating the genetic approach used to knockout of Per2 mice. B Schematic overview of experimental design for the wildtype and Per2 −/− mouse model treated with OVA or PBS. C - E qPCR analysis and Western blot analysis the expression of PER2 in wildtype and Per2 −/− mouse lung tissues ( n = 3). F Non-invasive assessment of pulmonary function in mice after treatment with methacholine ( n = 5). G - K Total BALF cells, differential inflammatory cell counts of the lung sections were performed by Giemsa staining ( n = 5). L Representative images of H&E, PAS, and Masson staining of lung tissues from mice, original magnification 200 ×. Scale bars, 50 μm. M – O Inflammation scores based on H&E staining, PAS score based on PAS staining, and Masson’ Trichrome stain of lung tissues from mice ( n = 5). All data are expressed as mean ± SEM. Statistical significance was determined using unpaired Student’s t-test for (C, E); two-way repeated ANOVA followed by turkey’s comparison test for ( F ); and one-way ANOVA followed by Tukey’s post-hoc test for ( G - K , M - O ). AHR, airway hyperresponsiveness. * P < 0.05, ** P < 0.01, *** P < 0.001, ns, not significant

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: Knock-Out, Western Blot, Expressing, Staining, Comparison

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: PER2 deficiency aggravates airway remodeling via Wnt/β-catenin signaling pathway in vivo. A - E Representative Western blot images and statistical plots of protein quantitative analysis of N-cadherin, E-cadherin, MMP-9, and Vimentin in wildtype and Per2 −/− in lung tissues of mice ( n = 3). F - H qPCR analysis of the expression of Tgfb1 , Mmp2 and Muc5ac in lung tissues of mice ( n = 3). I - L Representative immunohistochemical images and quantitative estimation of E-cadherin, N-cadherin, and α-SMA in lung tissues of mice, original magnification 200 ×. Scale bars, 50 μm. All data are expressed as mean ± SEM. Statistical significance was determined using one-way ANOVA followed by Tukey’s post-hoc test for ( B - H , J - L ). * P < 0.05, ** P < 0.01, *** P < 0.001, ns, not significant

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: In Vivo, Western Blot, Expressing, Immunohistochemical staining

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: PER2 deficiency aggravates airway remodeling via Wnt signaling pathway in vivo. A - B A KEGG enrichment analysis in the lung tissues of WT-OVA compared to Per2 −/− -OVA. C - D Gene expression of TGF-β signaling pathway and EMT was assessed by Gene set enrichment analysis (GSEA). E - I Representative Western blotting images and statistical plots of protein quantitative analysis of β-catenin, GSK-3β, and p-GSK-3β in protein extracted from the lung tissues of mice ( n = 3). All data are expressed as mean ± SEM. Statistical significance was determined using one-way ANOVA followed by Tukey’s post-hoc test for (F-I). * P < 0.05, ** P < 0.01, *** P < 0.001, ns, not significant

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: In Vivo, Gene Expression, Western Blot

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: PER2 overexpression attenuates TGF-β1-induced EMT and migration in bronchial epithelial cells. A - B Representative Western blotting images and quantitative estimation of PER2 expression in TGF-β1-induced BEAS-2B cells for different doses was analyzed. C qPCR analysis of the PER2 expression in TGF-β1(10 ng/mL)-induced BEAS-2B cells. D - E Representative IF staining of PER2 in BEAS-2B cells stimulated with TGF-β1, original magnification 200 ×. Scale bars, 100 μm. F - H The expression of PER2 was studied by using qPCR and Western blot analysis. I - L Representative Western blotting images and quantitative estimation of N-cadherin, E-cadherin, α-SMA expression in TGF-β1-induced BEAS-2B cells. M - O qPCR analysis of the expression of MMP2 , SNAI1 and SNAI2 in BEAS-2B cells. P - Q Representative images and quantitative analysis of cell migration rates showing changes in BEAS-2B cell migration as assessed by scratch assays, original magnification 100 ×. Scale bars, 100 μm. R - S Representative images and quantitative analysis of cell migration rates showing changes in BEAS-2B cell migration as assessed by transwell assays, original magnification 100 ×. Scale bars, 100 μm. Statistical significance was determined using one-way ANOVA followed by Tukey’s post-hoc test for ( B , F , H , J - O , Q , S ), and unpaired Student’s t-test for ( C , E ). All data are expressed as mean ± SEM ( n = 3). * P < 0.05, ** P < 0.01, *** P < 0.001, ns, not significant

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: Over Expression, Migration, Western Blot, Expressing, Staining

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: PER2 suppresses TGF-β1-induced EMT by inhibiting Wnt/β-catenin signaling activation. A A volcano plot showed Wnt signaling pathway and other circadian related genes (red, upregulated genes; blue, downregulated genes). B A The heatmap of differentially expressed genes. Results were based on 3 samples of RNA sequencing. C A KEGG enrichment analysis in the cultured BEAS-2B cells of PER2 OE + TGF-β1 compared to Vector + TGF-β1. D - H Representative Western blotting images and Statistical plots of protein quantitative analysis of β-catenin, GSK-3β, and p-GSK-3β in TGF-β1-induced BEAS-2B cells. I - L Representative Western blotting images and quantitative analysis of the expression levels of the N-cadherin, β-catenin, α-SMA following intervention with the β-catenin agonist SKL2001. M - O The effect of SKL2001 on the protein content of β-catenin was detected by Western blot in the cytosol and nuclear. P - Q The protein expression of β-catenin in nuclear and its quantification were identified by Immunofluorescence, original magnification 200 ×. Scale bars, 100 μm. All data are expressed as mean ± SEM ( n = 3). Statistical significance was determined using one-way ANOVA followed by Tukey’s post-hoc test for ( E - H , J - L , N , O , Q ). * P < 0.05, ** P < 0.01, *** P < 0.001, ns, not significant

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: Activation Assay, RNA Sequencing, Cell Culture, Plasmid Preparation, Western Blot, Expressing, Immunofluorescence

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: Immunofluorescence results of PER2 suppresses TGF-β1-induced EMT by inhibiting Wnt/β-catenin signaling activation. A - C The immunofluorescence analysis of N-cadherin, E-cadherin, and α-SMA expression after expression of PER2, original magnification 200 ×. Scale bars, 100 μm. D - F The relative fluorescence intensity of N-cadherin, E-cadherin, and α-SMA. All data are expressed as mean ± SEM ( n = 3). Statistical significance was determined using one-way ANOVA followed by Tukey’s post-hoc test for ( D - F ). * P < 0.05, ** P < 0.01, *** P < 0.001, ns, not significant

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: Immunofluorescence, Activation Assay, Expressing, Fluorescence

Journal: Respiratory Research

Article Title: The circadian clock component PER2 deficiency aggravates airway epithelial remodeling through Wnt/β-catenin signaling pathway

doi: 10.1186/s12931-026-03522-8

Figure Lengend Snippet: Melatonin exerts its protective effects against EMT of asthma by upregulating the PER2. A , Chemical Structure of melatonin is shown. B , Establishment of mouse models of OVA-induced asthma and melatonin treatment. C , The mRNA expression level of Per2 after treatment with melatonin in the lung tissues of mice was analyzed by using qPCR ( n = 3). D-E , Representative Western blotting images and quantitative estimation of PER2 after treatment with melatonin in wildtype and Per2 -/- mouse lung tissues ( n = 3). F , Non-invasive assessment of pulmonary function in mice after treatment with melatonin ( n = 5). G-K , Total BALF cells and differential inflammatory cell counts were performed by Giemsa staining ( n = 5). L-N , Representative lung sections and semiquantitative analysis of peri-airway inflammatory infiltration, and goblet cell hyperplasia in OVA-exposed mice after melatonin administration, original magnification×100. Scale bars, 50 μm. O-R , Representative Western blotting images and quantitative estimation of N-cadherin, E-cadherin, and Vimentin in the lung tissues of mice ( n = 3). All data are expressed as mean ± SEM. Statistical significance was determined using one-way ANOVA followed by Tukey’s post-hoc test for ( C , E , G - K , M , N , P - R ); and two-way repeated ANOVA followed by turkey’s comparison test for (F). * P < 0.05, ** P < 0.01, *** P < 0.001, ns, not significant

Article Snippet: After blocking with 5% BSA at 37 °C for 30 min, sections were incubated overnight at 4 °C with anti-PER2 antibody (1:100, NB100-125, Novus) and anti-SCGB1A1/CC10 antibody (1:50, 26,909–1-AP, Proteintech).

Techniques: Expressing, Western Blot, Staining, Comparison

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: (A–C) C57BL/6 mice housed under intense light (IL; 10,000 lux, L:D 14:10 h) for 3,5, or 7 days were subjected to 60 min of in situ myocardial ischemia followed by 2 h reperfusion at ZT3 (9 a.m.) and compared with mice housed under standard room light (RL; 200 lux, L:D 14:10h,7 days) (mean ± SD; n = 6; ANOVA with Tukey’s multiple comparison test). (A) Infarct size measurements. (B) Parallel measurements of serum troponin-I by ELISA (mean ± SD; n = 6; ANOVA with Tukey’s multiple comparison test). (C) Representative images of infarcts. (D–F) Wheel running measurements during 7 days of RL or IL housing in C57BL/6J mice (L, light phase; D, dark phase; n = 6; Student’s t test). (D) Wheel running activity graphs. (E) Distance walked. (F) Circadian amplitude. (G) Cardiac PER2 luciferase activity indicating protein in mice after RL or IL for 7 days (mean ± SD; n = 4; all IL versus RL p < 0.05 via ANOVA with Tukey’s multiple comparison test). (H–J) Wheel running during 7 days of RL or IL housing in C57BL/6J and Per2 −/− mice (n = 5–6; ANOVA with Tukey’s multiple comparison test). (H) Distance walked. (I) Circadian amplitude. (J) Wheel running activity graphs. (K and L) Immunoblot and quantification for PER2 protein in seeing or enucleated (blind) C57BL/6J mice after 7 days of RL or IL at ZT3 (mean ± SD; n = 5; Student’s t test). (K) Immunoblot. (L) Protein quantification. (M) Troponin-I serum levels in seeing or blind C57BL/6J mice housed under RL conditions followed by 60 min ischemia and 2 h reperfusion at ZT3 or ZT15 (mean ± SD; n = 4; ANOVA with Tukey’s multiple comparison test). (N) Wheel running measurements during 7 days of RL or IL housing in blind C57BL/6J mice (mean ± SD; n = 4; Student’s t test). See also .

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: In Situ, Comparison, Enzyme-linked Immunosorbent Assay, Activity Assay, Luciferase, Western Blot

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: (A) Infarct sizes in C57BL/6J mice that were housed under intense light (IL; 10,000 lux, L:D 14:10 h) for 7 days and subjected to 60 min of in situ myocardial ischemia followed by 2 h reperfusion at ZT3 or ZT15 (mean ± SD; n = 6; Student’s t test). (B–D) C57BL/6J mice exposed to voluntary wheel running for 1 versus 2 weeks. Shown are infarct sizes after 60 min of myocardial ischemia and 2 h reperfusion at ZT3 (B) or circadian amplitude (C) and distance walked measurements in relation to infarct sizes (D, mean ± SD; n = 6; Student’s t test). (E–H) Wheel running measurements during or infarct size studies after 2 weeks of wheel running at ZT3 in C57BL/6J or Per2 −/− mice (mean ± SD; n = 5; Student’s t test). (E) Distance walked. (F) Circadian amplitude. (G and H) Infarct size measurements (G) and one representative infarct size staining and one wheel running activity recording are shown (H). (I and J) Adenosine (I) or cAMP (J) levels in heart tissue from C57BL/6J or Per2 −/− mice at ZT3 after 7 days of room light (RL; 200 lux, L:D 14:10 h) or intense light (IL; 10,000 lux, L:D 14:10 h) housing (mean ± SD; n = 5; ANOVA with Tukey’s multiple comparison test). (K) Cardiac U- 13 C-glucose-1,6-bisphosphate levels at ZT3 from C57BL/6J mice that were housed under RL or IL for 7 days (mean ± SD; n = 4; Student’s t test). (L and M) Phosphofructokinase (PFK) activity in both heart tissue (L) and plasma samples (M) from C57BL/6J or Per2 −/− mice at ZT3 after 7 days of RL or IL housing (mean ± SD; n = 4–5; ANOVA with Tukey’s multiple comparison test). (N) HIF1A-hypoxia response element (HRE) binding was determined at ZT3, ZT9, ZT15, and ZT21 (mean ± SD; n = 5; *p < 0.05 for ZT21 versus ZT3 in RL- and IL-housed mice via Student’s t test). (O) C57BL/6J or Per2 −/− mice housed under IL for 7 days before 60 min myocardial ischemia and 2 h reperfusion at ZT3 (mean ± SD; n = 5; Student’s t test). (P) Representative infarct staining.

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: In Situ, Staining, Activity Assay, Comparison, Clinical Proteomics, Binding Assay

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: (A) Whole-genome array from C57BL/6J or Per2 −/− heart tissue after 7 days of intense light (IL; 10,000 lux, L:D 14:10 h) or standard room light (RL; 200 lux, L:D 14:10 h) housing at ZT3 (n = 3 per group, total of 12 arrays). Top light-regulated pathways are shown. (B) Validation of transcript levels of the top light and PER2-dependent gene (ANGPTL-4) identified by whole-genome array (mean ± SD; n = 4–5; Student’s t test). (C) Per2 mRNA transcript levels from endothelial cells isolated from endothelial-specific PER2-deficient ( Per2 loxP/loxP -VE-Cadherin-Cre) or control (VE-Cadherin-Cre) hearts (mean ± SD; n = 3; Student’s t test). (D and E) Infarct sizes (D) or serum troponin-I (E) in Per2 loxP/loxP -VE-Cadherin-Cre) or VE-Cadherin-Cre mice housed under RL or IL conditions for 7 days followed by 60 min of in situ myocardial ischemia and 2 h reperfusion at ZT3 (mean ± SD; n = 5; ANOVA with Tukey’s multiple comparison test). (F) Representative infarct staining. (G–I) Vascular leakage of Evans blue dye in C57BL/6J (G and H) or Per2 loxP/loxP -VE-Cadherin-Cre (I) after 60 min of in situ myocardial ischemia and 2 h reperfusion at ZT3 following 7 days of RL or IL housing (mean ± SD; n = 5; Student’s t test for G and ANOVA with Tukey’s multiple comparison test for I). (G) Vascular leakage quantification in C57BL/6J. (H) Representative Evans blue staining in C57BL/6J. (I) Per2loxP/loxP-VE-Cadherin-Cre. (J) Vascular leakage of Evans blue dye in Ador-a2b −/− after 60 min of in situ myocardial ischemia and 2 h reperfusion at ZT3 following 7 days of RL or IL housing (mean ± SD; n = 5; Student’s t test). (K) ChIP assay for HIF1A binding to the promoter region of Angptl4 in C57BL/6J following 7 days of RL or IL housing (mean ± SD; n = 3; Student’s t test). See also and .

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: Biomarker Discovery, Isolation, Control, In Situ, Comparison, Staining, Binding Assay

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: HMEC-1 or stable lentiviral-mediated PER2KD and Scr control HMEC-1 were synchronized and exposed to 24 h of normoxia (Nx) or 1% hypoxia (Hx). In a subset of experiments, synchronized stable lentiviral-mediated HIF1AKD and Scr HMEC-1 were exposed to Nx or Hx. (A and B) Affinity purification-mass spectrometry-based proteomics screen for PER2 protein interactions in normoxic and hypoxic HMEC-1. (A) Number of PER2 proteins regulated. (B) Pathways analysis using Ingenuity. (C and D) Coimmunoprecipitation for PER2 in hypoxic or normoxic HMEC-1 against isocitrate dehydrogenase (IDH) 2, succinyl coenzyme A (CoA) ligase (SUCLG) 1, and aconitase (ACO) 2 (C), and vice versa (D). One representative blot of three is displayed. (E) Subcellular compartment analysis of PER2 during normoxia or hypoxia (C, cytoplasm; N, nucleus; M, mitochondria; compartment-specific loading controls: tubulin alpha 1a (TUBA1A) for cytoplasm, TATA-box binding protein (TBP) for nucleus, and voltage-dependent anion channel 1 (VDAC1) for mitochondria). (F) Translocation of PER2 into the mitochondria during hypoxia (scale bar, 20 μm). (G–I) TCA cycle enzyme activities of IDH (G), SUCLG (H), and ACO (I) from stable lentiviral-mediated PER2KD and Scr control HMEC-1 during hypoxia (mean ± SD; n = 3; Student’s t test). (J) Carbon dioxide evolution rate (CDER), as a surrogate for TCA cycle function, in PER2KD or Scr HMEC-1 measured by a mitochondrial stress test using a Seahorse XF24 FluxPak assay (mean ± SD; n = 5; Student’s t test). (K–M) SIRT3 transcript (K and L) or protein (M) levels from stable lentiviral-mediated PER2KD and Scr (K and M, upper panel) or stable lentiviral-mediated HIF1AKD and Scr (L and M, lower panel) control HMEC-1 (mean ± SD; n = 3; ANOVA with Tukey’s multiple comparison test). See also – .

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: Control, Affinity Purification, Mass Spectrometry, Binding Assay, Translocation Assay, Comparison

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: (A–D) Oxygen consumption rates (OCRs) in PER2KD or Scr HMEC-1. Quantification of basal respiration, maximum achievable respiration, and ATP production are shown (mean ± SD; n = 5; Student’s t test). (A) Seahorse mitochondrial stress test. (B) Basal respiration. (C) Maximal respiration. (D) ATP production. (E) COX4.2 transcript levels in PER2KD or Scr HMEC-1 after 24 h of Nx or 1% Hx treatment (mean ± SD; n = 6; ANOVA with Tukey’s multiple comparison test). (F) Complex IV enzyme activity in Per2 −/− or C57BL/6 mouse hearts subjected to 45 min of ischemia (mean ± SD; n = 4; ANOVA with Tukey’s multiple comparison test). (G) Cardiac Cox42 mRNA levels at ZT3, ZT9, ZT15, and ZT21 in C57BL/6 mice after 7 days of room light (RL) or intense light (IL) housing (mean ± SD; n = 5; #p < 0.05 for ZT3 IL versus ZT3 in RL-housed mice via two-way ANOVA with Sidak’s multiple comparison test). (H) MitoTracker red CMXRos staining of PER2KD or Scr HMEC-1 at baseline. One representative image of five is shown (scale bar, 20 μm). (I) Quantification of the mitochondrial membrane potential probe JC-1 (mean ± SD; n = 6; ANOVA with Tukey’s multiple comparison test). (J–M) 13 C metabolites from supernatants of PER2KD or Scr HMEC-1 following 24 h of Nx or 1 % Hx treatment. Data are presented as the percentage of total metabolites present (mean ± SD; n = 3; ANOVA with Tukey’s multiple comparison test). (J) 13 C fructose-6-phosphate. (K) 13 C α-ketoglutarate. (L) 13 C 6-phosphogluconate. (M) 13 C palmitic acid. (N) Permeability assay in PER2KD or Scr HMEC-1 during 24 h of 1% hypoxia (mean ± SD; n = 5; two-way ANOVA with Tukey’s multiple comparison test). Note that permeability increases after prolonged hypoxia exposure of endothelial cells due to morphological changes. (O) CLDN1 (claudin-1) transcript levels in PER2KD or Scr HMEC-1 after 4 h of Nx or 1% Hx treatment (mean ± SD; n = 3; ANOVA with Tukey’s multiple comparison test). (P) Cardiac Cldn1 mRNA was determined at ZT3, ZT9, ZT15, and ZT21 in C57BL/6 mice after 7 days of RL or IL treatment (mean ± SD; n = 5; #p < 0.05 for ZT3 IL versus ZT3 in RL-housed mice via two-way ANOVA with Sidak’s multiple comparison test). See also and .

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: Comparison, Activity Assay, Staining, Membrane, Permeability

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: (A) Study design and verification of melanopsin overexpression by immunoblot. pCMV6 is the empty vector control, and OPN4-pCMV6 is the plasmid containing the gene encoding melanopsin (n = 3). (B–H) cAMP (B), pCREB levels (C), PER2 transcript (D) seahorse glycolytic stress test (E), glycolytic capacity (F), seahorse mitochondrial stress test (G), and maximum achievable respiration (H) after light-sensing cells were exposed to intense light (mean ± SD; n = 6–10; Student’s t test). (I) Schematic model.

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: Over Expression, Western Blot, Plasmid Preparation, Control

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: (A) Protocol for intense light exposure experiments in healthy human volunteers. 20 healthy volunteers (11 female and 6 male, age range between 21 and 44 years) were exposed to intense light (10,000 lux) from 8:30–9:00 a.m. on 5 consecutive days. (B and C) PER2 protein levels from buccal tissue (B) or plasma samples (C) at 9 a.m. during 5 days of intense light exposure assessed by immunoblot or ELISA, respectively (mean ± SD; n = 6; ANOVA with Tukey’s multiple comparison test). (D) Effect of room light versus intense light on human plasma melatonin levels (mean ± SD; n = 3–6; ANOVA with Tukey’s multiple comparison test). (E) Longitudinal monitoring of human plasma melatonin levels during 5 days of intense light exposure at 9 a.m. (mean ± SD; n = 3–6; ANOVA with Tukey’s multiple comparison test). (F) Human plasma phosphofructokinase (PFK) activity during 5 days of intense light exposure at 9 a.m. (mean ± SD; n = 3–6; ANOVA with Tukey’s multiple comparison test). (G) Human plasma PFK activity after 5 days of intense light exposure at 9 p.m. (mean ± SD; n = 3; Student’s t test). (H) Human plasma triglyceride levels during 5 days of intense light exposure at 9 a.m. (mean ± SD; n = 8; ANOVA with Tukey’s multiple comparison test). (I–K) Targeted metabolomics using mass spectrometry on human plasma samples from healthy volunteers exposed to intense light therapy for 5 days. (I) Pathway analysis. Key metabolites of glycolysis (pyruvate) or the TCA cycle (succinate, K) are shown for day 3 and day 5 of intense light therapy (mean ± SD; n = 3; ANOVA with Tukey’s multiple comparison test). (L–P) Actigraphy data using a validated accelerometer (Actiwatch 2). Shown are the wake after sleep onset (WASO) episodes (L), sleep efficiency (M), day activity (N), circadian amplitude (O) (mean ± SD; n = 6; Student’s t test), and one representative actigraphy recording from one healthy volunteer (P) before and during intense light therapy (synchronized sleep phases [turquoise bar] during intense light exposure [red square]). C, control subjects before light exposure; IL, intense light. See also and .

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: Clinical Proteomics, Western Blot, Enzyme-linked Immunosorbent Assay, Comparison, Activity Assay, Mass Spectrometry, Control

Journal: Cell reports

Article Title: Intense Light-Mediated Circadian Cardioprotection via Transcriptional Reprogramming of the Endothelium

doi: 10.1016/j.celrep.2019.07.020

Figure Lengend Snippet: KEY RESOURCE TABLE

Article Snippet: The primary antibodies used were rabbit polyclonal PER2 (Novus Biologicals, NB100-125, Littleton CO, or Abcam, ab64460, Cambridge, MA), mouse monoclonal actin (Ab-1) (JLA20, Calbiochem, Diego, CA,), rabbit polyclonal IDH2 (Novus Biologicals, NBP2-22166, Littleton CO), rabbit polyclonal SUCLG1 (Novus Biologicals, NBP1089489, Littleton CO), rabbit polyclonal ACO2 (Novus Biologicals, H00000050-D01P, Littleton CO), rabbit polyclonal SIRT3 (Abcam, ab86671, Cambridge, MA), anti-alpha Tubulin antibody (Abcam, ab7291, Cambridge, MA), Anti-VDAC1 / Porin antibody (Abcam, ab15895, Cambridge, MA), Anti-TATA binding protein (TBP) antibody (Abcam, ab51841, Cambridge, MA), mouse monoclonal β-ACTIN (Cell Signaling Technologies, 8H10D10, Danvers, MA), and mouse monoclonal anti-DDK (FLAG) (OriGene Technologies, TA50011-100, Rockville, MD).

Techniques: Binding Assay, Virus, Variant Assay, Recombinant, Protein Extraction, Extraction, Isolation, Cell Culture, Membrane, Transfection, Enzyme-linked Immunosorbent Assay, Reporter Assay, Activity Assay, Colorimetric Assay, Transcription Factor Assay, Bicinchoninic Acid Protein Assay, Chromatin Immunoprecipitation, Qubit Protein Assay, SYBR Green Assay, LDH Cytotoxicity Assay, Microarray, Luciferase, Generated, shRNA, Sequencing, Control, Software

Journal: Scientific Reports

Article Title: Circadian rhythms in pediatric high-grade gliomas may contribute to treatment efficacy

doi: 10.1038/s41598-025-17461-9

Figure Lengend Snippet: Circadian gene expression varies between pHGGs and pLGGs. ( A ) Interleaved scatter plots represent bulk RNA sequencing (rsem FPKM) analysis comparing circadian gene expression across pediatric low-grade glioma (pLGG, n = 426, blue) and pediatric high-grade glioma (pHGG, n = 211, red) samples from the Pediatric Brain Tumor Atlas (PBTA, Provisional). Data are expressed as mean ± SEM, and expression values are log₂-transformed FPKM counts. Statistical analysis was performed using a two-way ANOVA with Benjamini and Hochberg false discovery rate (FDR) to determine differences in gene expression between pHGGs and pLGGs. Each dot represents an individual tumor sample. Asterisks indicate statistically significant differences (* P < 0.05), while “NS” denotes non-significant comparisons. ( B ) Western blots of four representative pediatric high-grade glioma tissue samples (left) and four pediatric low-grade glioma tissue samples (right) comparing BMAL1, CLOCK, PER2, and REV-ERBα protein expression (sections of separate blots displayed; full blots are shown in Supplementary Figure S3). Of note, tissue sample HGG 3 is derived from the same patient as the STN-49 cell line. ( C ) Normalization to β-ACTIN control reveals BMAL1 ( p < 0.05, 1.57 vs. 0.98) and CLOCK ( p < 0.05, 0.98 vs. 0.39) protein expression is higher in high-grade gliomas compared to low-grade gliomas, REV-ERBα is lower ( p < 0.05, 0.12 vs. 0.55), and there are no significant differences in PER2 protein expression. Data are expressed as mean ± SEM.

Article Snippet: The following antibodies were used for immunoblotting: BMAL1 Rabbit mAb (A4714; ABclonal Technology, Woburn, MA), NR1D1 mAb (MA5-20772; Invitrogen), CLOCK anti-Rabbit (NB100-126; Novus Biologicals), PER2 anti-rabbit (NBP2-93587; Novus Biologicals), Beta-actin-HRP (MA5-15739-HRP; Invitrogen), anti-rabbit IgG HRP (7074; Cell Signaling Technology), and Goat anti-Mouse IgG Secondary Antibody, HRP (62-6520; Invitrogen).

Techniques: Gene Expression, RNA Sequencing, Expressing, Transformation Assay, Western Blot, Derivative Assay, Control

Journal: Cell metabolism

Article Title: Autophagy regulates the liver clock and glucose metabolism by degrading CRY1

doi: 10.1016/j.cmet.2018.05.023

Figure Lengend Snippet: KEY RESOURCES TABLE

Article Snippet: Rabbit polyclonal anti-PER2 , Origene , Cat# TA337016.

Techniques: Virus, Plasmid Preparation, Recombinant, Reverse Transcription, SYBR Green Assay, Electron Microscopy, Extraction, Software, Real-time Polymerase Chain Reaction, Microscopy