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I met Shelby Hinte on X, née Twitter, back when it was still a fun place to make jokes and connect with other writers. We started exchanging work, and the first manuscript of hers I read was an early version of Howling Women (Leftover Books
Sabine’s story seems more important to me than ever: when she leaves her husband and lover in Oakland with no particular destination or goal but escape
she’s forced to confront her past—specifically
the lasting legacy of her stepfather’s sexual abuse
which she has never so much as uttered before she meets the intriguingly named Howling Woman in Yu
In an era when it seems like most of them will never face consequences—legal
or reputational—Sabine’s burning anger speaks directly to mine
it was a pleasure to talk with Shelby about this novel’s genesis and evolution
Rebecca van LaerHowling Woman is such an interesting character
someone who could be cringe: her mystical name
Shelby HinteI spent a lot of my life around women like her—women with a confidence that enamored me but also made me uncomfortable
These women are in their late sixties and grew up calling themselves hippies and have lived in yurts and traveled in vans
And they sometimes say things that are inappropriate or culturally appropriative
I’ve never read Women Who Run With the Wolves
but these women have read and talked about it
And Sabine “knows better,” in terms of cultural appropriation
but not in terms of how to live with herself
You chose to have Sabine narrate the story from a point after her probation
when she has complete knowledge of Howling Woman's life
What made you decide on this narrative perspective rather than telling it from an earlier point in the timeline
SHSabine telling the story while awaiting trial was one of the last iterations of the book
but the book's always been interested in the same question: How did this happen
and she often writes about the impossibility of truth —how all we have is our own very specific truth
How does anyone make sense of what's happened to them
I wanted to explore a character who’s done something seemingly reckless and thinks: I'm an educated person; I had a somewhat normal life; and yet I did this thing that
RVLYou mentioned that this was a late iteration
Can you talk about what inspired that revision
there wasn’t anything at stake; she was just telling this story
If I want someone to read and I'm competing with the internet and television
I didn't want it to read like a television show
And I always thought of this book as loosely adjacent to a thriller
what if I just lean into that a little bit more
And so I upped the stakes by having her actually shoot the man
they are thinking constantly about how to tell their story
So there was a natural progression once I realized that that was what she had to be up against
you’ve taught me to always ask what my characters want
And how did you make sense of Sabine’s competing wants in this book
He taught us to plot around a character’s desires
Another good friend of mine asked in early drafts of this
but it also created a way to make the book character-driven
I created a character who desires something to an extreme degree
the desire for things causes me the most suffering
Desire has gotten me into trouble many times in the past
I feel like a huge part of my life is just trying to right-size my desire
What Sabine wants changes throughout the book
but what she originally wants is to escape the past—to be free of the human condition
which is that things happen to you and they stick with you forever
I think she just wants to be loved for exactly who she is
but it’s also thinking about plot in such smart ways
Sabine is often talking about plot: the inciting incident
SH This goes back to my question: How do stories get formed
I personally love stories that are aware of themselves as stories
and I liked the idea of her life being the story
but it seems to me like a wild act to ever commit your life to the page—I feel so different all the time about the past
I think we like to rewrite the past to fit new narratives we’ve gotten
I could explore how she tells her life story at this particular moment
RVL Let’s talk a little bit more about life
You're writing about childhood sexual abuse
What kind of self-care did you practice while inhabiting this narrative voice for so long
I feel pretty sick all the time when I'm writing
is because it truly feels like such an altered state
getting to inhabit another character feels like a way to be outside of myself
Self-care is something I’m maybe not very good at
but there is something really powerful about writing into a question that you care very deeply about
but even though it is very much a work of fiction
There are things in the book that mirror parts of my experience
I can take a question or experience I had and bring it to a new edge and play out like the worst thing that could have happened
or I could play out the best thing that could happen
It’s really healing to get to be in control of what happens to a character
No one else gets to be in charge of what happens here
RVL I’m so interested in how you’re talking about writing as an escape
and also what you said earlier about trying to right-size your desire
Do you think writing is a way for you to right-size it or to explore it and experience it in a safe container
SH It's a way to right-size my desire in my actual life
but what I've told my students when we talk about this is that the fun thing about fiction is that you can throw your characters into the scenarios that you've pondered without facing the repercussions
When I'm thinking of a character who has the same question I do
I can let it explode and go in any direction
If they come undone and unravel their whole life
and I also don't have to pick up the pieces in my personal life
RVL I’ve already asked you about what Sabine wants
The greatest compliment that I've had is I read it in a day
and takes me out of the real world for a little bit
So if I can give anyone the gift of getting to be consumed by art
this is a book about the harm men cause to women and the way women are not believed
I wondered if it was too extreme or unbelievable
I don't personally know a woman who hasn't experienced some form of assault
the woman gets to have agency and be a little crazy and take revenge
And I hope my readers take something away from that
Rebecca van Laer is a writer based in the Hudson Valley
Her work appears in the New England Review
Metrics details
Overcoming this barrier for long-term treatment success is difficult because the molecular mechanisms underpinning this phenomenon remain largely unknown
we show that both human and mouse adipose tissues retain cellular transcriptional changes after appreciable weight loss
we find persistent obesity-induced alterations in the epigenome of mouse adipocytes that negatively affect their function and response to metabolic stimuli
Mice carrying this obesogenic memory show accelerated rebound weight gain
and the epigenetic memory can explain future transcriptional deregulation in adipocytes in response to further high-fat diet feeding
our findings indicate the existence of an obesogenic memory
largely on the basis of stable epigenetic changes
in mouse adipocytes and probably other cell types
These changes seem to prime cells for pathological responses in an obesogenic environment
contributing to the problematic ‘yo-yo’ effect often seen with dieting
Targeting these changes in the future could improve long-term weight management and health outcomes
These reports might be confounded by variations in cell type composition
which are poorly characterized in the AT during WL
and therefore serve foremost as indicators of cellular epigenetic memory
it remains unresolved whether individual cells retain a metabolic memory and whether it is conferred through epigenetic mechanisms
we set out to address this by first performing single-nucleus RNA sequencing (snRNA-seq) of AT from individuals living with obesity before and after significant WL
confirming the presence of retained transcriptional changes
by characterizing the epigenome of mouse adipocytes
which revealed the long-term persistence of an epigenetic obesogenic memory
Source Data
pointing to potential dysfunction in the AT after WL
Experimental setup of the WL study in AdipoERCre x NuTRAP mice
Biotinylated nuclei and GFP-tagged ribosomes are isolated from frozen tissue
ATAC–seq (nuclei) and TRAP–seq (ribosomes)
PCA of translatome (TRAP–seq) of labelled adipocytes from C
Each dot represents an individual biological replicate
MOFA plots showing the sample clustering along latent Factors 1 and 2 (left) and Factor 1 value distribution (right) across labelled adipocytes
Each dot corresponds to one biological replicate
For each replicate all six modalities are represented in one dot
Percentage of variance explained by each MOFA factor across one of six modalities
Dynamics of differentially H3K4me3-marked promoters (y axis) from H to HC
Dynamics of differentially H3K27me3-marked promoters (y axis) from H to HC
H3K27me3 (middle) and H3K27ac (right) at selected promoters of genes and the log2FC of TRAP–seq from comparisons against controls for the same genes
Distribution of normalized reads of H3K4me3 and H3K27me3 at the Cyp2e1 and Icam1 loci across conditions
Correlation coefficient R (Pearson) of quantified peaks of H3K4me1 and H3K4me3 against a hypothetical healthy control (n = 2–3 each) with s.d
PCA plots of quantified adipocyte-specific enhancers as marked by H3K4me1
Dynamics of differentially H3K4me1-marked enhancers (y axis) from H to HC (left) and HH to HHC (right)
H3K27ac status of genes linked to newly emerged enhancers marked by H3K4me1 (from c) in different conditions identified by the presence of an H3K27ac peak associated to the gene
Top (significant) pathway terms for genes linked to newly emerged acetylated enhancers for H and HC (left) and HH and HHC (right) on the basis of WikiPathways database (Fisher’s exact test
adjusted P < 0.05 by the Benjamini–Hochberg method for correction)
Proportion of down- and upregulated memory DEGs from TRAP–seq that can be explained by one or more epigenetic modality in HC (n = 13; n = 72) and HHC (n = 7; n = 36)
these results strongly suggest the presence of stable cellular
epigenetic and transcriptional memory in mouse adipocytes that persists after WL
Each dot represents an individual biological replicate of a pool of three mice
HC and CC_s mice were put on HFD for 4 weeks
Each dot indicates an individual biological replicate from two experiments
Proportion of up- and downregulated DEGs in HCH adipocytes that can be explained by DEG status at HC time point or transcriptional memory
Normalized expression of selected DEGs in HCH adipocytes that were recovered in HC but were still differentially marked by one or more epigenetic modalities (Wilcoxon rank-sum test
adjusted P < 0.05 by the Bonferroni correction method; FC > ±0.5)
Distribution of normalized reads of H3K4me3
H3K27ac and H3K4me1 of HC and CC_s adipocytes at loci of Tmsbx4 and Gpam
Proportion of up- and downregulated DEGs in HCH adipocytes that can be explained by an epigenetic memory
Significance was calculated between age-matched controls and experimental groups
f and g was calculated using two-tailed Mann–Whitney tests
Significance for c was calculated using unpaired
Krieger and Yekutieli post-hoc test for multiple comparisons
these findings suggest that a persistent epigenetic memory
including local changes of hPTM deposition
contributes to the altered transcriptional response in adipocytes in the ‘yo-yo’ model of dieting and primes adipocytes for pathological responses to further HFD feeding
thus contributing to the pathophysiology of rebound obesity in mice
It is possible that other epigenetic modifications
we consistently observed retained transcriptional differences after significant WL in AT cells after sleeve gastrectomy (MTSS and LTSS studies)
which resulted in a complete return to a non-obese or lean state
The aforementioned alterations and the degree of WL achieved between individuals and studies are confounders that limit the direct comparability of our mouse and human data
The rapid WL achieved by BaS may even reduce or modify putative cellular memory in the human AT
Owing to the current lack of methods to isolate pure adipocyte nuclei from frozen human tissue
we could not perform the corresponding epigenetic analyses in human samples
it stands to reason that obesity-induced transcriptional (and cellular) changes in humans are also mediated through epigenetic mechanisms that can persist after WL in the AT and contribute to human (patho)physiology
also establish an epigenetic memory of obesity and contribute to the observed systemic weight regain effect
indicating that at least these treatments do not induce stable
Whether this is also the case for other agonists remains to be investigated
Further studies are needed to elucidate whether these treatments could erase or diminish an obesogenic memory better than other non-surgery-based WL strategies
No statistical methods were used to predetermine sample size
and the investigators were not blinded to allocation during experiments and outcome assessment
Human AT biopsies were obtained from three independent studies: MTSS
Samples of healthy individuals who were not obese were collected during routine elective surgeries such as herniotomies
explorative laparoscopies and cholecystectomies at the same hospitals
The study was approved by the Ethics Committee of the University of Leipzig under approval number 159-12–21052012 and was performed in agreement with the Declaration of Helsinki
Samples from healthy donors that were not obese were collected during routine elective surgeries (herniotomies
The study was approved by the Ethics Committee of the University of Leipzig under approval number 159-12–21052012 and performed in agreement with the Declaration of Helsinki
snap-frozen in liquid nitrogen and stored at −80 °C
The study was conducted in accordance with the Declaration of Helsinki and approved by the Ethics Committee of the Karolinska Institute
Stockholm (approval number 2011/1002-31/1)
Homozygous NuTRAP and AdipoERCre mice were bred to generate AdipoERCre x NuTRAP mice
AdipoERCre x NuTRAP mice were kept on HFD or chow diet for 12 or 25 weeks before tissue harvest or diet switch
The HFD used contained 60% (kcal%) fat (diet no
Provimi Kliba); the low-fat chow diet used contained 10% (kcal%) fat (diet no
During the WL period both experimental groups received chow diet (diet no
All animal experiments were approved by the Cantonal Veterinary Office
The 4–5-week-old AdipoERCre x NuTRAP mice were gavaged two times with 1 mg of tamoxifen dissolved in corn oil
Tamoxifen was washed out for 2 weeks before starting HFD
Mice were fasted for 6 h during dark phase before administration of 1 g of glucose per kg body weight by intraperitoneal injection
Blood was collected from the tail vein at 0
90 and 120 min and blood glucose concentrations were measured using an Accu-Check Aviva glucometer
Mice were fasted for 6 h during dark phase before administration of 1 U per kg body weight of human insulin (insulin Actrapid HM
Novo Nordisk) by intraperitoneal injection
90 and 120 min and blood glucose concentrations were measured using a Accu-Check Aviva glucometer
Measurements were obtained from one 8-cage and one 16-cage Promethion Core Behavioral System that were in the same room
Mice were habituated to the system for 36 h before measurements were started
Mice were fasted for 6 h during dark phase
Live mouse body composition was measured with a magnetic resonance imaging technique (EchoMRI 130
Fat and lean mass were analysed using EchoMRI 14 software
EDTA plasma was isolated from fasted blood samples (fasting 6 h)
Insulin was measured with Ultra Sensitive Mouse Insulin ELISA Kit (Crystal Chem
EDTA plasma (50 µl) was thawed on ice and used in a custom U-PLEX assay (Meso Scale Discovery) according to the manufacturer’s instructions
A Mesoscale SI 2400 was used to read the plate
50 mg of frozen liver was homogenized in 1 ml of isopropanol
lysed for 1 h at 4 °C and centrifuged for 10 min at 2,000g at 4 °C
The supernatant was transferred into a new tube and stored at −80 °C until use
Triglyceride levels were measured by mixing 200 µl of reagent R (Monlab
SR-41031) and 5 µl of sample or Cfas calibrator dilutions (Roche
then incubating for 10 min while shaking at room temperature and measuring optical density at 505 nm (OD505) with a plate reader (BioTek Gen5 Microplate Reader)
AT was minced and digested at 37 °C while shaking in collagenase buffer (25 mM NaHCO3
2.5% BSA; pH 7.4) using 2 mg of collagenase type II (Sigma-Aldrich
and for ingAT digestion continued for 15 min whereas epiAT was processed immediately
An equal volume of growth medium (DMEM (Gibco
15140-122)) was added and digested tissue was centrifuged for 4 min at 300g
and the floating fraction was transferred into a new Falcon tube and kept at 37 °C
The SVF was resuspended in 5 ml of erythrocyte lysis buffer (154 mM NH4Cl
filtered through a 40 µM mesh filter and centrifuged for 5 min
The SVF was resuspended in growth medium and counted
A total of 10,000 cells were plated into one well of a collagen-coated (Sigma-Aldrich
C3867) 96-well plate and kept in culture until they reached confluency
medium was changed to induction medium (DMEM
0.5 mM 3-isobutyl-1-methylxanthin (Sigma-Aldrich
After 48 h medium was changed to maintenance medium (DMEM
The SVF was cultured as described and controls were either kept in growth medium or only maintenance medium without induction
LZ-PT-7009) reagent was used according to the manufacturer’s instructions and read with a plate reader (BioTek Gen5 Microplate Reader)
Packed adipocytes (30 µl) were seeded onto one membrane and kept in inverted culture for 48 h in maintenance medium (DMEM-F12 (Gibco
adipocytes were washed and serum and glucose starved overnight in KREBBS-Ringer buffer (120 mM NaCl
pH 7.4) and 2.5% fat-free BSA (Sigma-Aldrich
Glucose uptake from primary adipocytes was measured using the Glucose Uptake-Glo Assay Kit (Promega
J1341) according to the manufacturer’s instructions
Adipocytes were preincubated with 5 nM insulin for 15 min before 2-deoxy-d-glucose was added at 1 mM final concentration
Protein concentration was measured using a Pierce 660 nm Protein Assay Kit (Thermo Fisher
22662) and the Ionic Detergent Compatibility Reagent (Thermo Fisher
Both assays were read with a plate reader (BioTek Gen5 Microplate Reader)
Starved adipocytes were incubated with 5 nM BODIPY-palmitate (Thermo Fisher
D3821) in the presence of 10 nM insulin for 1 h
adipocytes were washed twice and lysed in 200 µl of RIPA buffer
100 µl of lysate was used to measure BODIPY signal
Diluted lysate was used to measure protein concentration using a DC Protein Assay Kit II (Bio-Rad Laboratories
fixed in 4% PBS-buffered formalin for 72 h at 4 °C and stored in PBS at 4 °C
tissues were sent to the pathology service centre at Instituto Murciano de Investigación Biosanitaria Virgen de la Arrixaca for sectioning
Tissues from two independent experiments were sent for sectioning
Five to ten images were taken of each section belonging to a biological replicate (n = 4)
Nuclei were isolated from snap-frozen epiAT in ice-cold Nuclei Extraction Buffer (Miltenyi
130-128-024) supplemented with 0.2 U µl−1 recombinant RNase Inhibitor (Takara
2313) and 1× cOmplete EDTA-free Protease Inhibitor (Roche
5056489001) using the gentleMACS Octo Dissociator (Miltenyi
Nuclei were subsequently filtered through a 50 µm cell strainer (Sysmex
04-0042-2317) and washed two times in PBS-BSA (1% w/v) containing 0.2 U µl−1 RNase inhibitor
Nuclei were isolated from snap-frozen human AT (10–50 mg) in ice-cold Nuclei Extraction Buffer (Miltenyi
130-128-024) supplemented with 1 U µl−1 recombinant RNase Inhibitor (Takara
1× cOmplete EDTA-free Protease Inhibitor (Roche
5056489001) and 10 mM sodium butyrate using the gentleMACS Octo Dissociator (Miltenyi
The nuclei suspension was filtered through a 50 µm strainer
supplemented with PBS-BSA (1% w/v) containing 1× protease inhibitor and RNase inhibitor and centrifuged at 4 °C
The nuclei pellet was resuspended in 1 ml of PBS-BSA (1%
w/v) supplemented with RNase inhibitor (0.5 U µl−1) and 1× protease inhibitor and was transferred into a new 1.5 ml tube
Nuclei were counted using a haemocytometer and Trypan blue
concentration was adjusted to approximately 1,000 nuclei per µl and they were loaded onto a G-chip (10x Genomics
Single-cell gene expression libraries were prepared using the Chromium Next GEM Single Cell 3′ v3.1 kit (10x Genomics) according to the manufacturer’s instructions
two cycles were added to the complementary DNA amplification PCR
Libraries were pooled equimolecularly and sequenced in PE150 (paired-end 150) mode on a NovaSeq 6000 with about 40,000 reads per nucleus at Novogene or using a NovaSeqX at the Functional Genomics Center
Nuclei and ribosomes were isolated from snap-frozen epiAT from AdipoERCre x NuTRAP mice in ice-cold Nuclei Extraction Buffer (Miltenyi
The nuclei suspension was filtered through a 50 µm strainer and centrifuged at 4 °C
The supernatant was transferred into a new tube and supplemented with 2 mM dithiothreitol
01810) and 1 mg ml−1 sodium heparin (Sigma-Aldrich
w/v) supplemented with 0.2 U µl−1 RNase inhibitor
1× cOmplete EDTA-free Protease Inhibitor and 10 mM sodium butyrate and transferred into a new 1.5 ml tube
Nuclei were centrifuged and subsequently bound to Dynabeads MyOne Streptavidin C1 beads (Thermo Fisher
65002) for 30 min at 4 °C followed by three washes with PBS-BSA (1% w/v)
25 µl of GFP-Trap Magnetic Agarose Beads (ChromoTEK
gtma-20) were washed in 2 ml of polysome lysis buffer (50 mM TRIS-HCl pH 7.5
The supernatant was mixed with the beads and incubated at 4 °C on a rotator for 1–2 h
tubes were put on a magnetic stand and the supernatant was removed
The beads were washed three times with polysome lysis buffer supplemented with 2 mM dithiothreitol (Sigma-Aldrich
D0632-10G) and 1 mg ml−1 sodium heparin (VWR
ACRO411210010) and resuspended in 1 ml Trizol (Thermo Fisher
Trizol preserved samples were kept at −80 °C until RNA isolation
RNA was isolated by adding 200 µl of chloroform (Sigma-Aldrich
followed by shaking and centrifugation at 4 °C
The aqueous phase was transferred into a new tube and RNA was isolated and DNase treated with the RNA Clean and Concentrator-5 kit (Zymo Research
RNA libraries were prepared by performing reverse transcription and template switching using Maxima H Minus reverse transcriptase (Thermo Fisher
a template switch oligo and an oligodT primer to generate full-length cDNA
cDNA was amplified using the KAPA Hotstart 2x ReadyMix (Roche Diagnostics
1–3 ng of cDNA was tagmentated using 1.3 µg of Tn5 and amplified using KAPA HiFi plus dNTPs (Roche Diagnostics
07958846001) and the following PCR settings: 72 °C 5 min
Libraries were quantified using the KAPA library quantification kit (Roche Diagnostics
and sequenced in PE150 mode on a NovaSeq 6000 at Novogene
All buffers were supplemented with 1 x cOmplete EDTA-free Protease Inhibitor and 10 mM sodium butyrate
nuclei bound to beads were aliquoted into 96-well LoBind plates (Eppendorf
0030129547) and incubated with primary antibodies—anti-H3K4me3 (abcam
and the beads were resuspended in secondary antibody solution (guinea pig anti-rabbit IgG (antibodies-online
ABIN101961)) and incubated at room temperature
and transposition was performed at 37 °C and stopped using TAPS-Wash solution
Nuclei were lysed and pA-Tn5 decrosslinked using SDS-release solution
PCR was performed using KAPA HiFi plus dNTPs (Roche Diagnostics
07958846001) with the following PCR settings: 72 °C 5 min
Beads with nuclei were resuspended in ATAC–seq solution (10 mM TAPS pH 8.5
0.2 µg µl−1 transposase (Tn5)) and incubated at 37 °C for 30 min
100 µl of DNA binding buffer (Zymo Research
D4003-1) was added and samples were stored at −20 °C
DNA was extracted using Zymo DNA Clean and Concentrator-5 (Zymo Research
Library amplification was performed using KAPA HiFi plus dNTPs (Roche Diagnostics
Both ATAC–seq and CUT&Tag libraries were cleaned using SPRI beads
eluted in nuclease-free water and pooled equimolecularly after library quantification using the KAPA library quantification kit (Roche Diagnostics
Libraries were sequenced in PE150 mode on a NovaSeq 6000 at Novogene
we analysed tissue composition and removed nuclei belonging to donors in the case in which no nuclei were assigned as adipocytes (one case in NEFA) or more than 50% or nuclei were assigned as B cells (one case in MTSS; lean donor) after correspondence with surgeons
DEGs from obese and WL cells from mouse and human were overlayed
A DEG was considered restored if it was no longer deregulated in WL cells when compared with controls
we considered a DEG part of a transcriptional memory
Clusters identified as similar cell types (for example
three clusters of endothelial cells) were merged for DEG quantification but not differential expression analysis itself
only cell types for which we obtained at least 30 cells per donor were considered for the retention analysis
T cells were not included in differential expression analysis or transcriptional retention analysis
For integrated human adipocyte differential expression analysis quantification
with |log2FC| ≥ 1 and nominal P < 0.01 as cut-offs
The trained MOFA model represented data variability in terms of five latent factors
which were further explored and visualized
chromatin state fold enrichment was performed against genomic features and ENCODE candidate cis-regulatory elements
Enhancer states were selected on the basis of genomic localization and hPTM enrichment
an enhancer track was generated per condition and merged for differential analysis
Our raw peak count matrix was filtered for enhancer regions defined by chromHMM
and peaks around the TSS (±2,000 bp) were discarded
Linkage of putative enhancers to genes was done using the R package ChIPSeeker by selecting the closest gene (TSS or gene body) within 20,000 bp distance
Putative enhancers farther away than 20,000 from a TSS or gene body were not linked to any gene and were discarded from downstream GSEA
the raw filtered peak matrices were log-normalized using the R package EdgeR and Pearson’s correlation coefficient was computed using the cor function from the R package stats v.3.6.2
Differential analysis of the same hPTM between two conditions was performed using the R package EdgeR with nominal FDR < 0.05 and |log2FC| > 1 as cut-offs
Raw gene and promoter/enhancer-specific peak count matrices were log-normalized using the R package EdgeR
PCA of the normalized count matrices was performed using the prcomp function of R package stats v.3.6.2
For generation of heatmaps summarizing GSEA across cell types
significantly enriched terms were selected using the adjusted P value (<0.01) and the combined.score (enrichment score) was scaled and visualized
GraphPad Prism v.9.5.1 and Seqmonk v.1.48.1 were used to generate plots and Affinity Designer and Publisher were used to adjust plots for clarity (for example
GraphPad Prism v.9.5.1 was used to analyse physiological data from mice. Each dataset of physiological parameters was tested for normality using the Shapiro–Wilk test. On the basis of the results, parametric or non-parametric tests were used to compare experimental with age-matched control groups. Tests are indicated in figure legends and the Source Data
Further information on research design is available in the Nature Portfolio Reporting Summary linked to this article
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Henikoff, S., Janssens, D., Kaya-Okur, H. S., Henikoff, J. & Ahmad, K. CUT&Tag@home V1. protools.io https://www.protocols.io/view/cut-amp-tag-home-bd26i8he (2020)
H3K18 lactylation marks tissue-specific active enhancers
Integrated analysis of multimodal single-cell data
Doublet identification in single-cell sequencing data using scDblFinder
SoupX removes ambient RNA contamination from droplet-based single-cell RNA sequencing data
Perivascular mesenchymal cells control adipose-tissue macrophage accrual in obesity
A stromal cell population that inhibits adipogenesis in mammalian fat depots
Blanco-Carmona, E. Generating publication ready visualizations for single cell transcriptomics using SCpubr. Preprint at bioRxiv https://doi.org/10.1101/2022.02.28.482303 (2022)
Clustering trees: a visualization for evaluating clusterings at multiple resolutions
snRNA-seq reveals a subpopulation of adipocytes that regulates thermogenesis
Cellsnp-lite: an efficient tool for genotyping single cells
Vireo: Bayesian demultiplexing of pooled single-cell RNA-seq data without genotype reference
featureCounts: an efficient general purpose program for assigning sequence reads to genomic features
edgeR: a Bioconductor package for differential expression analysis of digital gene expression data
Peak calling by Sparse Enrichment Analysis for CUT&RUN chromatin profiling
The ENCODE blacklist: identification of problematic regions of the genome
ChIPseeker: an R/Bioconductor package for ChIP peak annotation
Andrews, S. SeqMonk mapped sequence analysis tool. Babraham Bioinformatics https://www.bioinformatics.babraham.ac.uk/projects/seqmonk/ (2022)
chromVAR: inferring transcription-factor-associated accessibility from single-cell epigenomic data
MOFA+: a statistical framework for comprehensive integration of multi-modal single-cell data
Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2
ChromHMM: automating chromatin-state discovery and characterization
Enrichr: interactive and collaborative HTML5 gene list enrichment analysis tool
Enrichr: a comprehensive gene set enrichment analysis web server 2016 update
Hinte, L. C. et al. Adipose tissue retains an epigenetic memory of obesity after weight loss. Zenodo https://doi.org/10.5281/zenodo.13870942 (2024)
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We thank all members of the von Meyenn group and the Wolfrum group for helpful discussions and support
von Wartburg for help with animal experiments and handling; J
de Sousa for help with computational pipelines; E
Dahlby for help with the metabolic cage measurements; E
Seelig for help with acquisition of human samples and scientific support; J
Waag for NuTRAP mice; and the Protein Production and Structure Core Facility at EPFL for the production and purification of pA-Tn5 and Tn5
This work was supported by ETH Zurich core funding (F.v.M.)
a European Research Council Starting Grant (no
the Basel Research Centre for Child Health (Multi-Investigator Project 2020 to F.v.M.)
the Deutsche Forschungsgemeinschaft (Project no
a European Research Council Synergy Grant (no
the Novo Nordisk Foundation (including the MeRIAD consortium grant no
the Knut and Alice Wallenberg’s Foundation (Wallenberg Clinical Scholar to M.R.)
the Strategic Research Program in Diabetes at Karolinska Institutet (M.R.) and the European Foundation for the Study of Diabetes (Future Leaders award to N.M.)
was funded by a postdoctoral grant from the Swedish Society for Medical Research
Open access funding provided by Swiss Federal Institute of Technology Zurich
Present address: Medical Oncology Department
Málaga Biomedical Research Institute (IBIMA)-CIMES-UMA
Present address: Helmholtz Institute for Metabolic
University of Leipzig and University Hospital Leipzig
Present address: Stem Cell Bio Regenerative Med Institute
Present address: Department of Bioengineering
These authors contributed equally: Laura C
Laboratory of Nutrition and Metabolic Epigenetics
Department of Health Sciences and Technology
Laboratory of Translational Nutrition Biology
performed mouse experiments and made libraries from mouse tissues
performed experiments with human samples and cell culture experiments
provided human AT samples and clinical data
collected additional metadata of human samples
supervised the human studies and human AT sample collections
interpreted the data and wrote the manuscript
All authors contributed to editing and reviewing the manuscript and approved its publication
received honoraria as a consultant and speaker from Amgen
received honoraria as a consultant and speaker from AstraZeneca
The other authors declare no competing interests
Nature thanks Katherine Gallagher, Naomi Habib, Alyssa Hasty and the other, anonymous, reviewer(s) for their contribution to the peer review of this work. Peer reviewer reports are available
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations
UMAP of 19,494 nuclei representing omAT pools from lean subjects (n = 5; 1 male
4 females) and paired omAT from T0 and T1 (n = 8 each; 2 males
b,c Proportion of retained transcriptional changes in highly abundant cell types of MTSS omAT
UMAP of 31,721 nuclei representing scAT pools from lean subjects (n = 8; 8 females) and paired scAT from T0 and T1 (n = 7 each; 7 females) from the NEFA study
e,f Proportion of retained transcriptional changes in highly abundant cell types of NEFA scAT
g-j Number of upregulated and downregulated DEGs per cell type obese donor scaled by column at T0 for omAT (left) and scAT (right) from MTSS
Number of persistently deregulated genes from T0 to T1 per cell type across AT pools from all studies
UMAP of 4,958 nuclei representing adipocytes from MTSS omAT and LTSS omAT (total lean n = 10; total T0/T1 n = 13)
UMAP of 13,231 nuclei representing adipocytes from NEFA scAT and LTSS scAT (total lean n = 13; total T0/T1 n = 12)
with adjusted p-value < 0.01 by the Bonferroni correction method and FC > ±0.5 was used for DEG identification in b
Cluster markers used for annotating cell clusters in human omAT of the MTSS (left) and LTSS (right) study
UMAP visualization representing omAT pools from the MTSS study (c) and LTSS study (d) coloured by predicted cell subtypes from the Emont et al
visceral AT dataset from Caucasian individuals
Feature plots showing reference mapping scores illustrating how well omAT dataset maps to the Emont et al
Relative cell type abundance in omAT per condition and tissue donor of the LTSS (e) and MTSS (f) study
Lines connecting dots indicate paired samples
Significance between T0 and T1 for e-f was calculated using paired multiple Wilcoxon tests with Benjamini
Krieger and Yekutieli post hoc test for multiple comparisons
Cluster markers used for annotating cell clusters in human scAT of the LTSS (left) and NEFA (right) study
UMAP visualization representing scAT pools from the LTSS study (c) and NEFA study (d) coloured by predicted cell subtypes from the Emont et al
subcutaneous AT dataset from Caucasian individuals
Feature plots showing reference mapping scores illustrating how well scAT dataset maps to the Emont et al
Relative cell type abundance in scAT per condition and tissue donor of the LTSS (e) and NEFA (f) study
Top (significant) persistently downregulated (memory) pathway terms in omental adipocytes of the MTSS (a) and LTSS (b) study based on Wikipathways database
Top (significant) persistently downregulated (memory) pathway terms in subcutaneous adipocytes of the LTSS (c) and NEFA (d) study based on Wikipathways database
Top (significant) persistently upregulated (memory) pathway terms in omental adipocytes of the MTSS (e) and LTSS (f) study based on Wikipathways database
Top (significant) persistently downregulated (memory) pathway terms in subcutaneous adipocytes of the LTSS (g) and NEFA (d) study based on Wikipathways database
Significance was calculated using Fisher’s exact test
with adjusted P-value < 0.05 by the Benjamini-Hochberg method for correction
each dot represents an biological replicate
Glucose tolerance tests (GTTs) and area of the curve (AOC) for GTTs; (n = 10 each)
Insulin tolerance tests (ITTs) and AOC for ITTs (n = 10 each)
GTTs and AOCs for GTTs; (n = 10 each from 2 independent experiments)
ITTs and AOC for ITTs (n = 10 each from 2 independent experiments)
Fasting blood glucose (CC_s&HC: n = 10 each
CCC&HHC: n = 20 each; from 2 independent experiments)
i,j Postprandial insulin and leptin levels
Cumulative food intake from HC and CC_s mice in the last 3 days of WL chow diet feeding
Energy expenditure of HC and CC_s mice in the last 3 days of WL chow diet feeding
Liver triglycerides (tg) per μg liver tissue (C&H: n = 6
Haematoxylin and eosin (HE) staining liver sections
Lean mass of HC and CC_s mice relative to lean mass measured at C and H timepoints of the same mice (right) (n = 19 each)
normalized to body weight (HH&CC_l: n = 6
Representative image of a histological and HE stained section of a whole epiAT depot from a HHC mouse
Haematoxylin and eosin (HE) staining of epiAT
Quantification of collagen content from Maison’s Trichome staining
Significance was calculated between age matched controls and experimental groups
z was calculated using two-tailed Mann-Whitney tests
two-tailed t-tests with Welch’s correction
Significance for p and r was calculated using unpaired
Cluster markers used to annotate cell clusters of mouse epiAT
UMAP visualization representing epiAT samples coloured by predicted cell subtypes from the Emont et al
Feature plots showing reference mapping scores illustrating how well this dataset maps to the Emont et al
UMAP of 16,567 nuclei representing macrophage subclusters
Number of upregulated (left) and downregulated (right) DEGs per cell type per comparison (H vs C
Proportion of retained transcriptional changes in different cell types
adjusted p-value < 0.05 by the Bonferroni correction method; FC > ±0.5)
Top (significant) persistently upregulated (memory) (c) and downregulated (d)
pathway terms in HC adipocytes based on Wikipathways database
Significant Wikipathways term enrichment scores related to persistently upregulated genes in HHC (f) and HC (g) per cell type
Significant Wikipathways term enrichment scores related to persistently downregulated genes in HHC (f) and HC (g) per cell type
Significance for c-h was calculated using Fisher’s exact test
Significant Wikipathways term enrichment scores related to genes associated with persistently differentially marked promoters by H3K27me3 (from e) or H3K4me3 (from e) in HHC adipocytes
Expression of genes encoding for epigenetic modifiers significantly deregulated either in H (*) or HH (#) adipocytes
adjusted p-value < 0.05 by the Bonferroni correction method; fold change (FC) > ±0.5)
None of the epigenetic modifiers are deregulated in HC or HHC adipocytes
Significance for f-g was calculated using Fisher’s exact test
candidate cis-regulatory elements as defined by ENCODE
not TSS-overlapping and with high DNase and CTCF signals only; DNase–H3K4me3
not TSS-overlapping and with high DNase and H3K4me3 signals only; dELS
TSS-distal with enhancer-like signatures; PLS
TSS-overlapping with promoter-like signatures; pELS
TSS-proximal with enhancer-like signatures
Correlation coefficient R (Pearson) of quantified peaks of H3K27ac against a hypothetical healthy control (n = 2-3 each) with s.d
ChromHMM analysis of the adipocyte hPTM profiles for conditions C
The colour scale corresponds to the emission parameter of each hPTM for each state
Fold enrichment of ChromHMM states from b and c for total genomic fraction coverage
PCA plot of quantified adipocyte specific enhancers from all conditions as marked by H3K4me1
PCA plot of quantified adipocyte specific enhancers as marked by H3K27ac
Top (significant) GO Cellular Component terms for genes linked to newly emerged and acetylated enhancers for H and HC (left) and HH and HHC (right)
adjusted p-value < 0.05 by the Benjamini-Hochberg method for correction)
cultured primary adipocytes from ingAT from CC_s and HC (left) and CCC and HHC (right) mice
Each dot represents an individual biological replicate of a pool of 3 mice
SVF stimulated with 10 nm insulin only (MM + Ins) and induced SVF with 10 nm insulin (IMM + Ins) 10 days after induction/no induction of differentiation from epiAT (b) and ingAT (c) SVF from CC_s
Every SVF pool was tested in all three conditions
GTT of HCH and CCH mice; blood glucose levels (n = 5 each)
ITT of CCH and HCH mice; blood glucose levels (n = 5 each)
Representative images of liver HE stained sections from CCH and HCH
Liver tg per μg liver tissue (C&H:n = 6 each
Pathological scoring of liver sections per group (n = 4 each)
UMAP of 15,665 nuclei representing epiAT pools (n = 5 pooled mice each) from CCH and HCH split by condition
Relative abundance of macrophage subclusters
Top significant pathway terms from upregulated (n) and downregulated (o) HCH DEGs that are explained by the epigenetic state in HC adipocytes based on Reactome database
was calculated using two-tailed Mann-Whitney tests
two-tailed Student’s t-tests with Welch’s correction and Benjamini
and Yekutieli correction for multiple testing
Significance for j was calculated using unpaired two-tailed Student’s t-tests with Welch’s correction
Gene counts and the number of unique molecular identifiers (UMIs) per condition of mouse epiAT samples
UMAP visualization representing integrated epiAT samples from the weight loss study (C
HCH) coloured by predicted cell subtypes from the Emont et al
Feature plots showing reference mapping scores illustrating how well these datasets maps to the Emont et al
gene counts and the number of UMIs per cell type from mouse epiAT samples
Gene counts and the number of UMIs per condition in the omAT samples from the MTSS (left)
LTSS (second left) and in scAT samples from the LTSS (second from right) and NEFA (right) study
Gene counts and the number of UMIs per donor in the omAT samples from the MTSS (b) and LTSS (c) study
Gene counts and the number of UMIs per donor in scAT samples from the LTSS (d) and NEFA (e) study
Gene counts and the number of UMIs per assigned cell type in the omAT samples from the MTSS (left) and LTSS (right) study
Gene counts and the number of UMIs per assigned cell type in the scAT samples from the LTSS (left) and NEFA (right) study
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DOI: https://doi.org/10.1038/s41586-024-08165-7
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weight loss as a result of lifestyle interventions
pharmacotherapy or bariatric surgery is often followed by weight regain
It is thought that cells retain an obesogenic memory that predisposes the body to return to the previous level of adiposity; however
the mechanisms that underlie this effect have been unclear
A new study has now demonstrated that adipocytes in humans and mice retain transcriptional changes after weight loss and that mouse adipocytes retain an epigenetic memory of obesity
retained transcriptional changes in human adipose tissue,” explain von Meyenn
The researchers observed similar changes in the samples from mice taken before and after weight loss
Obesity resulted in epigenetic changes at thousands of loci
and many of these changes persisted after weight loss
promoters that should be active remained silenced
suggesting that adipocytes truly retain an epigenetic memory of obesity,” say von Meyenn
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Hinte, L. C. et al. Adipose tissue retains an epigenetic memory of obesity after weight loss. Nature https://doi.org/10.1038/s41586-024-08165-7 (2024)
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DOI: https://doi.org/10.1038/s41574-024-01070-z
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Traditions run deep at Maryland’s HAT 50K
This article appeared in our August 2008 issue
We gladly forego the comfort of sofas and barstools and head for northeastern Maryland
to a park that’s spitting distance from where the Susquehanna River dumps into Chesapeake Bay
lubricate body parts and layer according to the latest forecast
We hit the trail en masse to give our legs and lungs the first real test of the year: the HAT 50K
The race’s 20th anniversary last March was my 15th time toeing the line
As I glance around in the brisk morning air
my mind fills with the usual abstractions and thoughts
familiar faces I haven’t seen since the fall
Jeff Hinte (the “H” in HAT) gives the start signal and Phil Anderson (the “A”) heads out across the field on his mountain bike
leading participants onto a short out-and-back road section that acts as a prelude to the trails (the “T”)
we pass the turn-of-the-century buildings that comprise the Steppingstone Museum
run through the Pavilion-turned-aid-station and disappear into the woods
first ventured into ultrarunning in the fall of 1987 at Maryland’s famous JFK 50 Mile
and as they extended the length of their training runs in preparation for a second attempt
they thought about staging a race of their own
“We had a great 7.5-mile loop in the Susquehanna State Park,” recalls Hinte
“and figured four laps was 3.8 miles longer than a marathon
so we gave it a shot.” Of the 46 who started the inaugural HAT run
The race doubled in size the following year and was led by its first international entrant
a Russian ultrarunner named Nail Bairmgalin on his first visit to the United States
the field reaches its 450-person limit well in advance
due largely to the co-director’s focus on making the experience about the runners
“We have strived to improve the quality of the event every year,” says Hinte
“and our incredible volunteers are the main reason we have been able to succeed.” Though the course has changed many times over the years (it currently consists of a short starting loop followed by two repeats of a long loop)
the aid stations are well placed and liberally stocked with everything from soup to nuts
“My philosophy is that if I wouldn’t use it
with hydration packs or courier bags replacing medals and plaques
you get everything you paid and then some back in good
useful gear.” With a number of top-10 finishes at the Wasatch and Vermont 100s and a victory at the original Old Dominion 100-miler
“Do you know who that is?” asks an unfamiliar voice from behind me
pointing to a red-shirted runner who is already building a lead
“Do you know who we are?” asks 42-year-old Ian Schouten
a fixture at the top of Pennsylvania’s trail-running scene for years and two-time HAT winner
is accompanied by 35-year-old Alex Barth of Chester Springs
Barth is another scary-good talent who has been winning everything in sight in recent years—including the Seneca Creek Trail Marathon
Nipmuck Marathon and Long Island Greenbelt Trail 50K—and his best still lies ahead
reminiscing about a winter training weekend that saw us fastpacking through snow-filled woods from dawn until dusk
“but he’ll be looking like that coyote we saw out there hiking by the end of the race
Dead meat is all I can say.” We burst into laughter at the words of bravado
it has taken its toll on Schouten as often as not
Described by Hinte as an event that is “both accessible to first-time ultrarunners and a challenging early season training run for veterans,” the HAT course seems deceptively runable early on
With the exception of a two-mile stretch of bitumen/gravel road in the middle of the big loop
The winding singletrack is some of Maryland’s most popular for mountain bikers
and it serves up challenging roller-coaster-like running
with continuous one- to three-hundred-foot losses and gains
the singletrack offers solid footing as it twists and turns through the deciduous forest
Schouten contemplates what the valley must have looked like before the 17th century settlers started timbering
“Imagine this whole area covered in trees of that size,” he marvels as we swing around the several-hundred-year-old sentinel and peer into its six-foot diameter trunk
We’re all ears.” We round a corner and spook a trio of whitetail deer
They crash off through the woods and soon blend into the landscape
Though there are stretches of green and the odd stand of emergent flowers when we hit the open fields
the open stretches of field could be mucky or frozen
sun-baked or buffeted by bone-chilling wind
The weather might be sub-freezing with snow squalls or provide a spike of unseasonable heat
and the pair of creek crossings can be waist deep or worse
but the conditions encourage foolhardiness
No single element of the course is especially daunting when viewed in isolation
they can brew up a Perfect Storm and lead to disaster for even the most experienced racer
HAT 2001 was our introductory meeting and laid the foundation for an intensely competitive
mutually supportive friendship that will last until we’re both long in the tooth
more technical races put on by Ron Horn of Pretzel City Sports fame in the rough trails near Reading
Schouten had become familiar with the HAT course while supervising a job site nearby
“I don’t know about you,” he said as we ran together in the opening miles
“but I’m thinking this pace is a little bit slow.”
Don’t let me hold you back though.” At the five-mile mark
as I hit the series of climbs where I eventually overtook Schouten in 2001 (he held on in two of our next three meetings)
I have just passed the Rock Run Grist Mill
erected in 1794 by a prosperous businessman from Baltimore
and am once again high above the Susquehanna
Distant sailboats on the blue water look like bright kites offset by the sky
Schouten and Barth have broken away in pursuit of Redpath and another runner
This is only the first lap and there’s still a long way to go
Thoughts shift to the depth of competitors that have taken on HAT over the years
National Team members and great local runners,” says Hinte
Cross Country Team member Terry Croyle won the sixth HAT
Leadville regular Christine Gibbons won the women’s division twice in a row in the late 1990s
Olympic Marathon Trials Qualifier Jacquie Merritt won in 2000
taking third place overall and becoming the only woman to break the four-hour mark.”
Philadelphia meteorologist Cecily Tynan won the race with an inspiring performance
“It was very difficult for me,” said the 32-year-old who had graced the cover of Runner’s World magazine the previous month
“because I’m not used to the trails.” I learned that Tynan was a nationally ranked duathlete with several marathon wins under her belt and wondered aloud if this was the start of an ultra career
National Team member and sub-four-minute miler Andy Downin was even more strident
Upon finishing the 2006 HAT in third place
he uttered four simple words: “You people are nuts!”
These are not the sentiments of most newcomers
the camaraderie or the people who put on the event
chances are you’ll finish with a smile on your face
You’ll forget the pain as soon as you dig into the veggie chili and chase it with a homemade cookie or 12
You’ll be hooked—and welcomed into the fold
“There’s just something special about this race compared to others I attend,” says Lou Shaffer
a perennial volunteer from Anderson’s hometown
Shaffer’s wife Bonnie finished her sixth HAT this year
captured the HAT spirit the best: “The whole day embodied the laid-back
happy-go-lucky personality of most trail runners
With all the great aspects of a small race
I was surprised to find out that it’s one of the largest in the country
“You bet,” said 63-year-old Anderson at the start of the race when I asked him if he was running this year
always an ultrarunner.” Despite suffering a stroke 11 months earlier
he hopped off the lead bike and joined the rest of us in celebrating the Equinox out on the trail
Barry Lewis is a Philadelphia-based writer with five HAT wins to his credit
my mind went boom: let's do a music festival and a film
We had some people turn us down – even personal friends like Stevie Wonder and Sly Stone – because we were all flying out together on our own plane with no extra equipment
They wanted to bring entourages and have their own planes
But it meant we ended up with the people who really wanted to be there
Everyone was easy to negotiate with, except James Brown
meaning he needed his sound system and back-ups
So he turned up at the airport with 32,000lb of extra equipment which almost stopped us going
I just closed my eyes and let them load it on
who had worked on planes and wanted to kill James
and people respected him anyway; once he got on stage you forgot you didn't like him
Hope She'll Be Happier – which he'd pulled out of nowhere at 3am
reflecting trouble he was going through with a girl – he has never sung since
Reading this on mobile? Click here to view video
The footage of the Zaire concert seemed to me so powerful that when I edited it into a documentary in 2008, I felt it should speak for itself. [Levy-Hinte had previously edited When We Were Kings (1996)
using footage shot by many of the same film-makers.] So the cinema verite quality the film has arose naturally out if that
Stewart had brought in these excellent film-makers
and many of the camera rolls struck me as near–perfect short films
In one scene, film-maker Paul Goldsmith happens upon a group of Kinshasa street musicians
He begins walking slowly down the road towards them until they gradually fill the frame
and the sound of their playing drifts over
Then he moves around them in a purely intuitive way
and then the drummer before he begins a flourish
Over the end credits we used footage by Albert Maysles
who tracked Brown back to his dressing room; there is not a wasted frame
He would have played to the camera all evening
but I think his people realised he was going to collapse
I also wanted to convey the prevalence of music in the everyday life of Zaire people
how it was in the rhythm of their movement
talking about how it felt like returning to his homeland and discovering
The pitch is the first step in an often very long process of making a movie
It’s what gets the money people to open
An asteroid is about to smash into the earth and only Bruce Willis can save us
wild comparisons are evoked to assure its marketability
It’s Terminator meets Harry Met Sally with a side of Toy Story
the idea is to go big and dramatic in just a few sentences because that’s all the time a writer has to convince a producer the project has merit
this summer’s breakout critical hit: “Something about siblings
one that ultimately does not lend itself to easy taglines — a lesbian couple whose teenage children decide to seek out their sperm donor father — it takes a particular producer
One who freely admits to being not particularly adept at knowing what qualities will make a hit
“The most important element for me is the dedication and creativity of the director,” said Jeffrey Levy-Hinte
“There is this person and I can really believe in what they’re trying to say.”
Many projects I’ve taken on I just couldn’t get made
It all comes down to finding financing.”
The Kids Are All Right could have just as easily been one of those other projects
whose credits as an editor and producer include Soul Power
believed in the vision of writer/director Lisa Cholodenko and her writing partner on this film
in the journey of his own life in the movie business
The pitch for that: Young twenty-something guy dissatisfied with his day job in commercial real estate ends up watching a lot of movies to escape
But then there is the dedication and creativity of the young man in question
or where I’d flee to in order to get away from the job
And then I just got it in my head that this is what I wanted to do
I began talking to people to see what this community was about
This was the mid-nineties and there was this new technology where they were using computers to edit for the first time
It created a great demand and very little supply.”
which focused on postproduction services such as editing and sound
the very successful documentary about Muhammad Ali and George Foreman’s classic fight in Zaire
After this commercial and critical success
offers to edit more documentaries flooded his office
to keep the story of this young man moving forward
How about one of the oldest story devices known
I had met Lisa [Cholodenko] earlier when I was editing
but I ran into her at the Barnes and Noble on Fifth Avenue and asked
‘What are you up to?’ and she said
Levy-Hinte’s first feature film production credit and it was Ms
Cholodenko’s first writing and directing credit
The two paired up again to make Laurel Canyon
a family drama starring Christian Bale and Frances McDormand
Almost immediately they began talking about their next movie together
“is she has this wonderful way of getting into the inside of her characters and really elaborating all these circumstances and scenarios they might be in.”
The Kids Are All Right was supposed to be a much darker film
“But then Lisa decided she didn’t want to go into the heart of darkness and instead tread on the periphery with some sunshine.”
the making of the movie did not receive the same warm rays of light
After about a year spent thinking about the story
Cholodenko teamed up with Stuart Blumberg and they created a treatment that had many of the elements the finished film has
mostly due to the precariousness of financing an independent movie
“People make grand gestures of what they intend to do,” said Mr
the stars began to align and the movie was about to be shot
Then a creation of another sort intervened: Ms
The actors had other projects to do and couldn’t wait around
“So we scheduled the shoot for her second trimester
always knowing it was completely insane.”
But the financing fell apart just weeks before shooting was to begin
The team decided to fold up their tents for awhile
even turning his attention back to documentaries
“We also used the time as an opportunity to rethink the script and it became a much more focused script
We were really able to answer the essential questions
Each individual character’s arc.”
She and Julianne Moore and the director began spending time with the story and inhabiting their characters’ lives
“Lisa would talk to them [Annette and Julianne] eliciting their views and listening to the cadence of their voices
Now the characters started to move in the direction of the people.”
just three weeks before shooting began in July of 2009
that they had created something that in all likelihood will end in a trip to the Oscars next year
“I was so focused on getting it made that it didn’t really occur to me that it could have this success
But then we’d have these screenings for small groups to talk about it
these people really like the movie.” And like it they did
The movie caused a bidding war at the Sundance Film Festival
eventually selling for five million dollars to Focus Features
It opened in theaters this summer and this week is still chugging along nicely at the box office
its course steered by a producer and creative team determined to make a film that meant something
the one about the young man who once escaped misery by watching movies only then to make producing movies his life’s work
he was in the process of leaving the industry
Looking back on his journey in the movie business
“First you get your foot run over by a roller skater
And then comes the car and then the bus and then one of those dump trucks that they use in strip-mining coal that go over you and then back up and go over you again.” Ouch
how about a Vineyard story to refuel the soul
Levy-Hinte first visited the Island in the early nineties but only briefly
a friend who knew of his interest in wooden boats and craftsmen in particular told him about Nat Benjamin
“So I came out a couple of times to see Nat
I’d visit him and photograph his shop
But I couldn’t find much in the way of a dramatic arc other than
Levy-Hinte started coming to the Island more frequently with his wife
who had been vacationing on the Vineyard for more than 20 years
They rented each summer until they discovered that the old Humphrey’s house in West Tisbury was for sale
where the turkey gobbler sandwich started and donuts big as your head fueled many a bicycler headed up-Island
But something there was crying out to be restored.” Another project to believe in
The house restoration took almost three years
They enjoyed their first summer there in 2009
just after shooting was completed on The Kids Are All Right
“I had gotten wind that Nat had started to work on his own boat
That’s something that could be interesting,” Mr
“That just might do it.” With the help of cinematographer Brian Dowley
he filmed the story of the making and launching of Charlotte
searching for the through line of the story of Charlotte
the one he was so intent on leaving and the film that has so many people talking
“What’s most important to me,” Mr
“is whether the film is meaningful and important to people and then they want other people to see it and it creates a following
is you need some measure of success that allows you to go back and do more.” In other words
Audiences can only hope that the success of this film inspires Mr
deciding whether to take their own leap of faith
Jeffrey Levy-Hinte will take questions after a screening on Wednesday at 8 p.m
part of the Martha’s Vineyard Film Festival
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speaking Friday during his company's first-quarter earnings call
not least because AOL has a long way to go
EMarketer has it ranked 11th in terms of global digital ad share
with just 0.71% of the market to Facebook's 9.07% and Google's 31.07% (see table below)
But fresh from announcing a 7.2% lift in revenue
Armstrong is determined the company will get there
And he gave two big reasons he thinks it's possible
where advertisers can plug in lots of different components of their spend — across AOL's platforms
and even TV — and view it all in one place
Armstrong said on the call that marketers were moving in this direction
They want to consolidate the number of vendors they work with when it comes to their digital advertising
He said "unsophisticated customers" had about "15 point solutions" whereas the more sophisticated customers were on a "migration path" to five or so systems
"We're building toward being one of the top three in this area," Armstrong said (Nos
Armstrong thinks AOL can differentiate itself from Google and Facebook by offering an "open" platform
where marketers can plug in their own components
Armstrong used the metaphor of computer network servers: "You don't buy big-box servers anymore
where you have to throw the whole server out — you can replace the different parts
Another way in which AOL (along with others
such as Yahoo) stands out from Facebook and Google is that it produces its own original content
which it says reaches 200 million monthly unique users; other popular news sites including TechCrunch and Engadget; and video programming such as "Park Bench" with Steve Buscemi and "Making A Scene" with James Franco
AOL says its top shows reach 10 million to 15 million viewers
which isn't bad considering its video offering is still new to the market
Armstrong describes this as its "barbell" strategy: "Premium" content
which is very different from the pure ad tech players it is fighting to take advertising share from
And it plans to invest about $25 million in video content over the coming year
who was also speaking on the first-quarter earnings call
It is the fastest-growing advertising sector in the US
AOL knows it can't take on the scale of Facebook and YouTube in this area (at least not yet)
but it hopes its two-pronged strategy can at least see it give the rest of the digital advertising sector a run for its money
there are already some signs of a revenue payoff
on the eve of the Martha’s Vineyard Book Festival
Kusama-Hinte and his crew filmed the documentary at Gannon & Benjamin Marine Railway in Vineyard Haven
he said the movie took over six years to complete
Although the documentary tracks the process of building one schooner
the movie captures an overall feel of working at a boat yard
Watch the video for the full interview with Mr
The MV Times comment policy requires first and last name for all comments
Newspaper claims it did not know images of Susanne Hinte it bought for £750 were stolen
The Sun is paying substantial damages to relatives of a woman whose lottery jackpot claim was rejected after it published revenge porn images of her on its website and in print
Susanne Hinte, who died last year, hit the headlines in early 2016 when she tried to claim a £33m prize with a ticket that she said had been damaged in the wash
Camelot rejected her claim but Hinte became a brief favourite with the tabloids
Later that year Hinte’s former friend Julie Howard sold topless pictures of Hinte to the Sun for £750
She went on to tell police she had done it to humiliate Hinte after they fell out
She obtained the pictures after Hinte borrowed her phone
which automatically copied the images Hinte took to a cloud storage service
The Sun published the photographs on its website and in print on Sunday 3 April 2016
In January 2017 Howard was found guilty of disclosing private sexual images with intent to cause distress
and received a fine and a six-week suspended sentence
She was convicted under a law the Sun had campaigned for in 2014
The newspaper claims it did not know the images it bought from her were stolen
Hinte died suddenly of a heart attack in August 2017
Her daughter, Natasha Douglas, pursued a claim against News Group Newspapers (NGN)
The claim was for the misuse of private information
copyright and data protection law and the distress that the article caused to Hinte
NGN has not admitted liability but has settled for a five-figure sum
When the settlement is read in open court on Thursday morning it will be the first statement ever to be read in open court in a privacy case for a deceased person
a partner and media and data privacy law specialist at JMW Solicitors
said: “This was a serious intrusion and misuse of Susanne’s private information
The Sun showed a complete disregard for her legal rights
As a direct result of its decision to publish the distasteful and unnecessary article
she and her children suffered considerable embarrassment
Douglas said: “I felt very strongly that it was important to continue this claim against the Sun on behalf of my late mother
not only because it’s what she would have wanted but also to hold them accountable and make other people aware that it’s totally unacceptable to intrude into someone’s life as they did in this case
“Their actions caused a huge amount of pain – revenge porn is an awful thing
This article was amended on 19 June 2018 to correct a date: the images were published on the website on 2 April 2016 and in print on Sunday 3 April
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This is the lottery ticket said to belong to Susanne Hinte
But this sorry-looking ticket may – the emphasis
has to be on the may – be worth a cool £33m
This is an image of a lottery ticket said to belong to Susanne Hinte
Family members have said that a ticket bought by Hinte at her local newsagent’s appearing to bear the winning numbers was found after being put through the wash in a pair of her jeans
has refused to comment on the case and said hundreds of people who believed they may have won had come forward
Image taken from social media believed to be that of Susanne Hinte
Photograph: FacebookHinte herself has kept a low profile at her flat since her name emerged in the media but the saga took at twist when this image purporting to be of the winning ticket surfaced
52 and 58 – do match those that were drawn out on Saturday 9 January but other identifying features used to verify that a ticket is genuine are missing
View image in fullscreenA National Lottery £66m jackpot specimen ticket
Photograph: David Davies/PA The date the ticket was purchased is missing from the top right corner
There appears to be a tear where this should be
which is reproduced on the top and bottom of the ticket
which is automatically generated when the ticket is purchased and gives the ticket holder the chance to win £1m
Two security barcodes – one at the top right
the other along the bottom – are incomplete
just under a fortnight after the draw was made
As is usual when it is trying to encourage a missing winner to come forward
Camelot announced the vague area where the winning unclaimed ticket had been bought
where around 50 shops sell lottery tickets
visited the city with a giant cheque made out to a “mystery millionaire” for £33,035,323
He said: “We want the people of Worcester to check their tickets
We want them to check down the side of sofas
on shelves – just anything that can help them find the winning ticket.”
lottery representatives in the area recommended four clean
friendly shops where the media could film and report from
run for the last 27 years by Natu and Hansa Patel
welcomed in – and charmed – the film crews
Camelot – or at least a “handful” of security experts – know exactly where and when the winning ticket was bought
word spread that the winning ticket was bought at the Patels’ store
Local reporters began digging for the winner
focusing on the area around Ambleside News
There was chatter on social media – and suddenly Hinte’s name popped out
She has not spoken or been pictured but her daughter, Natasha Douglas, told an agency reporter: “The ticket has been through the wash
She wanted to stay anonymous but obviously her name has got out through people talking on Facebook
When she found out she had the winning numbers she couldn’t breathe and she hasn’t slept since.”
It is no surprise she wanted to remain anonymous
In the past few days her life has been pored over
Friends said she was long separated from her second husband
revealed they were not divorced and wondered whether he could be in for a payday
Natu Patel with his wife, Hansa, at their shop, Ambleside News, in Worcester. Mr Patel said: ‘I remember her buying the ticket and I really hope it is the winning one.’ Photograph: Newsteam /SWNS GuardianCamelot continues to bat off requests to spill the beans about the claim
It refuses to say whether the winning ticket was bought at Ambleside
Natu Patel said Camelot had not asked him for his CCTV footage of his shop at the time the winning ticket was bought
The lottery operator said it had the discretion to pay prizes even if tickets were stolen
A spokesperson said: “If the player can provide sufficient evidence
we will investigate and consider the validity of the claim
Such evidence may include where and when the ticket was bought
how the numbers were chosen (eg lucky dip or chosen numbers)
how many lines played and other relevant information
If we subsequently determine that the claim is valid
we then have the discretion to pay the prize 180 days after the draw.”
But it said it had received hundreds of claims
“All of these are currently being considered on a case-by-case basis
and we will follow up with all claimants directly to advise them whether their claim will be investigated further
Given the volume and the fact that some claims may require further information from the players involved
“We will contact claimants as soon as possible
but obviously need to ensure we have thoroughly investigated all claims
We do not comment publicly on any ongoing claims and would only provide further information once a prize has been validated and paid out
this could not happen until 180 days after the draw at the earliest.”
Camelot warned people against making false claims
The spokesperson said: “With prizes of this size
it’s perfectly normal to receive lots of claims from people who genuinely think that they may have mislaid or thrown away what they believe was the winning ticket
and we are looking into all of these claims as part of our efforts to find the rightful ticket holder
if we believe that somebody has intentionally attempted to defraud the National Lottery
we reserve the right to take whatever action we consider is appropriate.”
One person who is remaining sanguine is Patel
“I remember her buying the ticket and I really hope it is the winning one,” he said
His trade in lottery tickets and in newspapers telling the story has certainly been boosted by the saga
“We are certainly seeing increased footfall
People are thinking this is a lucky shop.”It emerged later on Wednesday that Hinte is due to appear before magistrates charged with theft
The charge is not connected to the lottery ticket claim
spokesman for West Midlands Crown Prosecution Service
said: “We can confirm that Suzanna Hinte has been charged with two counts of theft
She will appear at Birmingham magistrates on 1 March.”
The CPS has a slightly different spelling to the one on the electoral roll at Hinte’s home
who claimed she put £33million winning ticket in washing machine found dead at homeThe 49-year-old is believed to have had a heart attack at her home on Monday afternoon
who tried to claim a £33 million lotto jackpot by alleging she had put the ticket in the washing machine
The 49-year-old is believed to have had a heart attack at her home on Monday afternoon
Last year the grandmother of four, known to friends as Sanne, contacted Camelot claiming a battered slip pulled out of her washing machine with no date or barcode was the lucky ticket.
She alleged she had won half the £66 million jackpot from the January 9th draw in 2016 after it was revealed the winning ticket was bought in her home town of Worcester.
German-born Sanne sent in the crumpled ticket but when officials confirmed the true winner had come forward with the real ticket her plan was exposed.
Susanne’s claim had been among several hundred when it was revealed the winner was from Worcester. But a source said most would have been dismissed immediately.
Neighbours described how police and an ambulance crew suddenly raced to Susanne Hinte's home yesterday afternoon.
She is believed to share the £175,000 semi detached home on a housing estate in Worcester with her son.
It is understood the German born gran-of four died as a result of heart failure having suffered with a heart condition for a number of years.
Her agent Barry Tomes said: "I was told last night but I was only able to verify it this morning that her son had found her at the house.
"I believe it was heart failure or a heart attack that killed her because she hadn't been very well for a number of years.
"People would accuse her of faking it - but obviously it was genuine and she couldn't walk more than a few metres without struggling.
"She had a lot of enemies but she was a very talented and creative woman and this is very sad news and my thoughts are with her family.
"Whatever she's been involved with in the past nobody deserves to go like that at such a young age."
One neighbour, who did not want to be named, said: "Its desperately sad. There was a lot of activity here last night and we knew it wasn't looking good.
"She had a lot of critics but we always found her to be a good neighbour and she was certainly quite a character."
A police spokesman confirmed: "Police were called by the ambulance service to an address on Borrowdale Drive, Worcester at 5:40pm yesterday (Monday, 14 August).
"A woman, aged 49, was declared dead at the scene. Her death is not being treated as suspicious and has been referred to the coroner.
West Midlands Ambulance spokesman said:"We can confirm we were called to reports of a woman in cardiac arrest at a property in Borrowdale Drive, Worcester, at 5.35pm yesterday (Mon).
"An ambulance and a paramedic officer attended the scene.
"Unfortunately, it quickly became apparent on arrival at the scene that nothing could be done to save the patient and she was confirmed deceased."
Last year she said: "I wanted to be dead
"I couldn't understand why all of a sudden I was hated by so many people
"I didn't do anything wrong."
In January her former friend was found guilty of a revenge porn offence after selling pictures of her topless and wearing lingerie to a national newspaper
Worcester magistrates' court was told Julie Howard received £750 for the images to get her back for hiding her hair curlers in a tumble dryer
Howard was given a suspended six-week jail term after she was found guilty of disclosing sexual pictures
Sanne had also been before the courts following the false claim
Last July she was cleared of stealing a woman's purse following a three day trial in Birmingham
And in April 2016 a charge that she stole an Xbox controller from a man she had met online was dropped when he declined to attend court
She lived at the house with son Brandon who earlier this year had lost a child
Sanne wrote on Facebook said: "Watching ur own child grieve over the loss of his own child has to be the most unbearable pain u can feel as a mother as nothing u say or do can take their pain away
"I love u so very much Brandon I am so very proud of you and Becky for being so very brave today which I no was the hardest day of your life
"And I'm so so glad I had the oppunity in meeting my little gran child while he lay fast asleep with Angel's watching over him keeping him safe u were perfect in every way."
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LOTTO gran Susanne Hinte has removed a Jacuzzi hot tub from her home just weeks after buying it with taxpayers' money
The jobless 48-year-old, who receives over £1,000 in benefits each month, bought the luxury bath unit in June and put it in her plush shed
She had told The Sun that she was using it to treat her enlarged heart – a condition that has prevented her from working
According to the Daily Star, the Worcester resident has now ditched the Jacuzzi after reading insulting comments about herself on the internet.
The German-born grandmother, who falsely tried to claim a £33 million Lotto jackpot in January and was later featured in a TV show, said: “I have got rid of my shed now. I have ripped it out.
“It is because what people were saying about me that made me so angry. It made me angry how people saw me [because of the shed].
“It was as if I made the shed just because I had the money [from benefits]. People out there thought it was because of that.”
I had to actually shut my Facebook down for a bit because it was so horrendous
a scumbag and that I was still trying to make money out of it [the lottery controversy]
but since February this year I have had to give up working because of my condition.”
Local residents and social media users were left outraged after it emerged Susanne had bought the hot tub in the summer
One disgusted neighbour said: “It is ridiculous really
I understand it has cost her around £1,000 and it is rather extravagant to be honest.”
One social media user wrote: “She has a bad heart but … she was alright when she was getting in and out of the pool whilst on holiday abroad I saw.”
Another wrote: “Give the money she's getting to someone who really deserves it.”
chief executive of the TaxPayers' Alliance
said: “Questions need to be asked about a system that allows claimants to pay for luxuries that hard-pressed taxpayers could never afford for themselves.”
In April of this year it was revealed she was to get a new £20,000 Vauxhall Astra to help her get around – also funded by the taxpayer
Do you have a story for The Sun Online news team
Email us at tips@the-sun.co.uk or call 0207 782 4368
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A gran who tried to claim a £33 million lotto jackpot by pretending the winning ticket had gone through the wash has died
Susanne Hinte, 49, is believed to have had a heart attack at her home on Monday afternoon
contacted Camelot after claiming she found the winning ticket in her jeans which had been damaged when it went through the wash
She alleged she had won half the £66 million jackpot from the January 9 draw in 2016 after it was revealed the winning ticket was bought in her home town of Worcester
But when the German-born gran sent officials the crumpled ticket
investigators from the security team soon realised something was amiss
Neighbours described how police and an ambulance crew suddenly raced to Susanne Hinte's home yesterday afternoon
She is believed to share the £175,000 semi-detached property
Ms Hinte provoked a storm of criticism by the false claim and later revealed she regretted the whole incident and felt suicidal
"I couldn't understand why all of a sudden I was hated by so many people
"I didn't do anything wrong."
In January her former friend was found guilty of a revenge porn offence after selling pictures of Susanne topless and wearing lingerie to a national newspaper
Worcester Magistrates' Court was told Julie Howard received £750 for the images to get her back for hiding her hair curlers in a tumble dryer
Ms Hinte was cleared of stealing a woman's purse following a three day trial in Birmingham
a charge relating to a stolen Xbox controller from a man Ms Hinte had met online was dropped when he declined to attend court
Ms Hinte wrote on Facebook: "Watching ur own child grieve over the loss of his own child has to be the most unbearable pain u can feel as a mother as nothing u say or do can take their pain away
"And I'm so so glad I had the oppunity in meeting my little gran child while he lay fast asleep with angels watching over him keeping him safe u were perfect in every way."
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Lotto gran Susanne Hinte is putting the drama of a failed bid to claim £33million behind her and turning her attention towards becoming a lingerie model
Susanne, 48, claimed her 'winning' ticket had become faded in the wash
with numbers and dates rubbed off after being left inside a jeans pocket
After Camelot launched an investigation into the claim
the gran's claim was revealed to be incorrect when the real winner came forward and scooped the massive jackpot
has been flooded with offers to model sexy designer lingerie for a range of boutique stores
The German gran-of-four seductively posed for cameras after trying on a grey underwear set made by Sipsey Lingerie in Solihull
Read more: Lotto gran Susanne Hinte sells her washing machine on eBay - but for how much?
turns to look down the lens with a white negligee draped over her shoulder
She was also offered a free bra fitting service as well as an array of designer lingerie by the luxury brand
Susanne said: "I have been overwhelmed by people trying to help me and the fact that someone would want to offer me designer lingerie is unbelievable."
The Lotto gran was also invited for a special consultation with bespoke bra designer Fitelle at their shop in Woolhampton
She spoke to owner and designer Debi Richens who created a mould of her breast after a personal consultation
Read more:Lotto gran who claimed she damaged £3m winning ticket in the wash reveals her swingers clubs shame
Susanne added: "Debi was so helpful and professional and really made me understand why it is important to wear the right bra
something I haven't done because I don't wear bras."
Bra expert Debi added: "It is a pleasure to help a woman who could benefit from having the support and shape of a custom made bra
In January Susanne tried to claim the joint Lotto prize after Camelot revealed the winning £33million ticket was bought in her home town of Worcester
She is also rumoured to have been in talks to appear on the next series of Celebrity Big Brother
All images are copyrighted by their respective copyright holders and/or producers/distributors
emma grittPublished: Invalid Date
chancer Susanne Hinte is still living a life of luxury
the 48-year-old puffs on a fag in a bubbling hot tub
dubbed 'Lotto Gran' after she was caught trying to scam Lottery chiefs to get her hands on an unclaimed £33 million jackpot
She'll feature in On Benefits: Life On The Dole
where cameras will follow her going about her daily business
gets a £500 monthly PIP plus around £600 in other benefits
She was also given a taxpayer-funded Vauxhall Astra GTC worth £20,000
A source close to Susanne told The Sun online: "Viewers will be granted an intimate look in to Susanne's daily life
"Since the lottery ticket incident she's gone from gran on benefits to a celebrity gran on benefits - with a lifestyle to match."
Cameras followed Susanne as she attended radio interviews
and will also be granted a look inside the garden shed she converted in to a relaxing hot tub space after enjoying one of the bubbling pools while staying at the five star hotel Malmaison hotel in Birmingham
Now it looks like fans could be seeing a lot more of Susanne - even though she reportedly turned down a £500 bid to star in an X-rated porn film
The source added: "If her appearance on Life On The Dole goes well it could open a lot of doors for Susanne
"There's no reason why this can't launch her as a reality TV star - there's certainly enough interest in her and she's got a real screen presence
people might be so impressed with the shed - which she did all the interior design for - she'll end up launching a lucrative new career doing that."
When she first opened the door to her luxury shed last month, she told The Sun it was her stint at the Malmaison that inspired her to have a hot tub at home.
“I am sure I will be criticised but at least my neighbours leave me alone now
Whatever I do I seem to get hated so I will close the door and relax in warm bliss."
Susanne certainly looks at ease as she enjoys a cigarette and a glass of fizz
she shows off the curves that saw her approached to be a lingerie model as her star soared in the days after the lottery ticket scandal
She made headlines when she claimed she had scooped the record prize, but said her ticket was missing vital parts after being damaged in the washing machine
On Benefits: Life On The Dole airs on Channel 5
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