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Rudolph, M. C.

Publications and source records attributed to Rudolph, M. C..

13 recordsLinked to original sources

A naturally evolved mutation in cavefish mc3r reshapes energy homeostasis across development in a nutrient-limited environment.

Food availability in natural habitats is often unpredictable and drives the selection of adaptive strategies in organisms to survive food scarcity. However, the genetic and molecular mechanisms that enable animals to survive under such nutritional extremes are poorly understood. Naturally adapted organisms therefore provide powerful genetic systems for uncovering the mechanisms that maintain energy homeostasis under nutritional extremes. Here, we leveraged the cavefish Astyanax mexicanus to identify a naturally occurring Thr166Met missense mutation in the melanocortin 3 receptor (mc3r) that reprograms energy homeostasis to facilitate adaptation to nutrient deprivation. To determine its functional consequences, we introduced the mutation into zebrafish mc3r using CRISPR/Cas9 genome editing. We show that adult mc3r mutants exhibit increased body size, adiposity, and hyperglycemia relative to wild-type animals, recapitulating key metabolic features of Tinaja cavefish and identifying mc3r as the first causal gene behind cavefish hyperglycemia. Investigations for early developmental consequences of the mutation revealed accelerated yolk utilization in mutant zebrafish larvae and strikingly elevated metabolic rates relative to WT, thus, uncovering a previously unrecognized role for mc3r in regulating energy expenditure. Importantly, the phenotype was also observed in Tinaja cavefish revealing a novel cavefish phenotype and establishing causal role for mc3r in elevated metabolic rate phenotype of cavefish. Together, these findings reveal that the Thr166Met variant produces distinct physiological consequences across life stages, enhanced energy storage and hyperglycemia in adulthood despite elevated larval energy expenditure. We propose that increased larval metabolism may support the energetic demands of enhanced foraging, whereas increased adiposity and blood glucose later in life provide energy reserves during periods of nutrient scarcity. More broadly, our study demonstrates how a single naturally evolved coding variant can reshape energy homeostasis across development and contribute to adaptation to nutrient-poor environments.

evolutionary biology↗

Disulfiram Protects Against Diet-Induced Obesity by Reprogramming Systemic Lipid Partitioning Independent of GSDMD

Obesity remains a major global health challenge with limited durable pharmacotherapies. Disulfiram (DSF), an FDA-approved drug reported to inhibit gasdermin D (GSDMD), has been proposed to improve metabolic outcomes through suppression of inflammasome signaling. Here, we demonstrate that GSDMD is dispensable for high-fat diet-induced obesity and insulin resistance, as neither genetic deletion nor antisense-mediated inhibition of GSDMD confers metabolic protection. In contrast, DSF robustly protects against obesity and IR through a GSDMD-independent mechanism. These effects are not attributable to reduced caloric intake but instead reflect a coordinated reprogramming of systemic lipid handling. Under steady-state conditions, DSF suppresses basal lipid oxidation while promoting fecal fatty acid excretion. In striking contrast, during acute lipid challenge, DSF enhances tissue lipid utilization and accelerates systemic clearance. Together, these findings overturn the prevailing inflammasome-centric model and establish context-dependent regulation of lipid partitioning--rather than inflammasome inhibition--as the primary mechanism underlying DSFs anti-obesity effects

pharmacology and toxicology↗

The Ets transcription factor ETV4 regulates FGF1-dependent proliferation and glycolysis in ER-positive breast cancer

Obesity is a risk factor for estrogen receptor (ER) positive breast cancer. Beyond body mass index, adult weight gain increases breast cancer risk. During weight gain, hypertrophic adipocytes produce fibroblast growth factor 1 (FGF1), which drives estrogen-independent growth of ER-positive tumors. Effects of FGF1 on breast cancer cells include elevated proliferation and enhanced glycolytic activity. We identified the Ets transcription factor ETV4 as a target of FGF1 treatment across multiple breast cancer cell lines. Our objective was to define the role of ETV4 in mediating the tumor-promotional effects of FGF1, to better understand how weight gain and obesity drive breast cancer risk and progression. Here, we determined that ETV4 directly associates with a poor prognosis for patients with ER-positive tumors and positively correlates with FGF1 levels in the context of obesity. We demonstrate that ETV4 is required to mediate the pro-tumorigenic effects of FGF1 on cell proliferation, glycolytic reprogramming, and tamoxifen sensitivity in vitro, and on tumor growth in the presence of estrogen in obese mice. In vitro, ETV4 overexpression enhances proliferation and metabolic activity, mimicking effects of FGF1 on breast cancer cells, but it is not sufficient to promote ER-positive tumor growth before or after estrogen deprivation in vivo in lean females. This study reveals a potentially novel mechanism through which weight gain, characterized by excess FGF1 production, drives the development of aggressive features in the prevalent ER-positive breast cancer subtype.

cancer biology↗

Tamoxifen Targets Wisp2 to Impair Subcutaneous Adipocyte Progenitor Self-Renewal and Adipogenic Differentiation

Breast cancer endocrine therapy, which systemically disrupts estrogen receptor signaling, increases type 2 diabetes (T2D) risk in some women. Sustained treatment with low-dose tamoxifen depletes subcutaneous adipocyte progenitors and promotes glucose intolerance and hepatic lipid deposition in obese female mice. Hyperplastic adipose tissue expansion, especially in subcutaneous depots, preserves metabolic health during a chronic positive energy balance by facilitating nutrient storage and attenuating inflammation. Adipocyte progenitors are renewed in part through Wnt signaling pathway activation, which is altered in women with obesity or T2D. Estrogen receptors are expressed in several adipose cell types, but the distinct actions of tamoxifen in adipocyte progenitors and the mechanisms that explain their depletion during endocrine therapy are not defined. The direct impact of tamoxifen was evaluated in subcutaneous adipose stromal cells from humans and adult mice. Self-renewal, proliferation, and differentiation were measured, and analyses of gene expression and progenitor or preadipocyte populations were performed. Mechanistic insight was gained from primary adipose stromal cells of obese female mice, in which the Wnt1 inducible signaling pathway protein 2 (Wisp2) was lost following endocrine therapy. Wisp2 gain and loss of function studies were carried out in adipose stromal cells to define the link between estrogen signaling and adipocyte progenitor maintenance. We found that tamoxifen treatment disrupts the protection of adipocyte progenitors by estrogen, mediated through suppression of Wisp2. These studies reveal potential metabolic effects of tamoxifen therapy that precede and could drive T2D development in breast cancer survivors.

cell biology↗

Lipodystrophy and recovery are mediated by the Wnt/lipogenesis axis during skin fibrosis.

Acquired lipodystrophy in the dermal white adipose tissue (DWAT) is a salient feature of skin fibrosis, and is followed by accumulation of extracellular matrix (ECM). Lipodystrophy syndromes, often associated with metabolic co-morbidities, are estimated to affect 1 in 20,000 people. We recently showed that fibrosis-associated lipodystrophy is dependent on sustained Wnt signaling, but the mechanism is unclear. Transcriptomic profiling of mature dermal adipocytes in vivo reveal that Wnt activation downregulates the de novo-lipogenesis (DNL) axis enzymes within 48 hours. We found that protein expression of Fatty Acid Synthase (FASN), a key DNL enzyme, is dependent on sustained Wnt activation in vitro and in vivo. In human systemic sclerosis, FASN mRNA is significantly downregulated during two years of disease. Remarkably, pharmacological inhibition of FASN enzyme during reversal from Wnt induced fibrosis impedes recovery of DWAT lipid content as well as ECM accumulation and topography. All together, we demonstrate that acquired lipodystrophy in the context of skin fibrosis is mediated by a new role of the Wnt-DNL axis. These findings underscore the importance of this pathway in lipodystrophy and fibrosis, opening new avenues for therapeutic targets in skin fibrosis.

cell biology↗

Maternal exercise during lactation reprograms obesity-related changes in mammary metabolism to optimize milk fatty acids and offspring energy expenditure

Maternal obesity alters breast milk composition in ways that may predispose infants to excess adiposity. While maternal exercise during lactation has been associated with favorable shifts in milk metabolites in humans, the mechanisms by which exercise remodels the mammary gland and milk lipid profile to influence offspring metabolism remain unclear. We developed a mouse model incorporating daily moderate treadmill exercise during lactation, indirect calorimetry, stable isotope tracer respirometry, and mammary epithelial cell (MEC) proteomics in lean (LN) and diet-induced obese (OB) dams. Maternal obesity broadly remodeled the MEC proteome, reducing enzymes of de novo fatty acid synthesis and altering lipid transport and oxidative pathways. These molecular adaptations corresponded to higher milk triglyceride content and shifts in fatty acid composition, including an elevated omega-6 to omega-3 fatty acid ratio. The exercise (EX) intervention during lactation reset MEC protein networks, enhancing translational and vesicle transport pathways while reducing fatty acid desaturation, relative to the sedentary (SED) group. In OB dams, exercise increased milk medium-chain fatty acid (MCFA) levels and partially corrected the n6/n3 FA ratio. Offspring nursed by OB-EX dams exhibited higher whole-body energy expenditure, increased fatty acid oxidation, and improved metabolic flexibility compared to litters consuming OB-SED milk. Together, maternal exercise during lactation remodels mammary metabolism and milk fatty acid composition in obese dams, enhancing neonatal lipid oxidation and energy expenditure. These findings highlight lactation as a modifiable window, wherein maternal activity influences milk composition and infant metabolic health. New and noteworthyMaternal obesity alters milk fatty acid composition, with consequences for infant metabolism. Exercise during lactation in obese dams remodeled the mammary epithelial cell proteome, increasing medium-chain fatty acids in milk and enhancing lipid oxidation and energy expenditure in offspring.

physiology↗

Diet-induced phospholipid remodeling dictates ferroptosis sensitivity and tumorigenesis in the pancreas

High-fat diet (HFD) intake has been linked to an increased risk of pancreatic ductal adenocarcinoma (PDAC), a lethal and therapy-resistant cancer. However, whether and how specific dietary fats drive cancer development remains unresolved. Leveraging an oncogenic Kras-driven mouse model that closely mimics human PDAC progression, we screened a dozen isocaloric HFDs differing solely in fat source and representing the diversity of human fat consumption. Unexpectedly, diets rich in oleic acid - a monounsaturated fatty acid (MUFA) typically associated with good health - markedly enhanced tumorigenesis. Conversely, diets high in polyunsaturated fatty acids (PUFAs) suppressed tumor progression. Relative dietary fatty acid saturation levels (PUFA/MUFA) governed pancreatic membrane phospholipid composition, lipid peroxidation, and ferroptosis sensitivity in mice, concordant with circulating PUFA/MUFA levels being linked to altered PDAC risk in humans. These findings directly implicate dietary unsaturated fatty acids in controlling ferroptosis susceptibility and tumorigenesis, supporting potential "precision nutrition" strategies for PDAC prevention.

cancer biology↗

Hypertrophic adipocytes increase extracellular vesicle-mediated lipid release and reprogram breast cancer cell metabolism

Primary adipocytes possess a dramatic capacity to expand and retract in volume, leading to high variability in cell size within and between individuals. Yet, how adipocyte size impacts cell function remains unclear as adipocyte size is not tunable with traditional experimental approaches, forcing previous work to rely on correlative studies. Here, we develop protocols to separate primary adipocytes from the same donor into large and small populations and maintain these size-sorted cells in culture. Using these methods, we perform transcriptomic, lipidomic, and functional analyses on large and small adipocytes across two orthogonal mouse models of obesity and validate our results with human clinical samples. Our findings indicate that changes to cell size, rather than global differences mediated by weight gain, drive the transcriptional response of primary adipocytes to obesity. Moreover, large adipocytes shift from a traditional, lipase-mediated mode of lipid release to a non-canonical, extracellular vesicle-mediated mechanism. In functional coculture studies, this change promotes lipid accumulation in neighboring breast cancer cells, increasing their migration and proliferation via enhanced tumor cell fatty acid oxidation. Consistent with our experimental data, human patients with large adipocytes present with greater rates of dyslipidemia and higher concentrations of fasting triglycerides, even when accounting for differences in body mass index. Collectively, our results provide direct evidence that large and small adipocytes from the same donor differ in gene expression, lipid composition, and function with implications for the management of adipose tissue-related pathologies such as breast cancer.

cell biology↗

Integrated omics analysis reveals human milk oligosaccharide biosynthesis programs in human lactocytes

Human milk oligosaccharides (HMOs) are integral to infant health. Yet, their complex biosynthesis pathways in the mammary gland during lactation remain under characterized. To address this knowledge gap, we performed integrated analyses of single-cell RNA-sequencing (scRNA-seq) datasets combined with select HMO concentration measures. We identify differential expression patterns of known HMO synthesis genes in epithelial subsets and nominate several candidate genes that vary with HMO concentration. Additionally, we identify novel gene patterns and transcription factors that may regulate the expression of HMO biosynthesis genes and the cellular pathways supporting HMO production. Finally, we demonstrate that co-expression of HMO synthesis genes and milk fat synthesis genes is limited, suggesting distinct epithelial cell subtypes may be responsible for the production of different milk components. Our study suggests that HMO synthesis may be achieved through cell type specialization within the lactocyte compartment.

systems biology↗

NR2F2 Reactivation in Early-life Adipocyte Stem-like Cells Rescues Adipocyte Mitochondrial Oxidation

In humans, perinatal exposure to an elevated omega-6 (n6) relative to omega-3 (n3) Fatty Acid (FA) ratio is associated with the likelihood of childhood obesity. In mice, we show perinatal exposure to excessive n6-FA programs neonatal Adipocyte Stem-like cells (ASCs) to differentiate into adipocytes with lower mitochondrial nutrient oxidation and a propensity for nutrient storage. Omega-6 FA exposure reduced fatty acid oxidation (FAO) capacity, coinciding with impaired induction of beige adipocyte regulatory factors PPAR{gamma}, PGC1, PRDM16, and UCP1. ASCs from n6-FA exposed pups formed adipocytes with increased lipogenic genes in vitro, consistent with an in vivo accelerated adipocyte hypertrophy, greater triacylglyceride accumulation, and increased % body fat. Conversely, n6-FA exposed pups had impaired whole animal 13C-palmitate oxidation. The metabolic nuclear receptor, NR2F2, was suppressed in ASCs by excess n6-FA intake preceding adipogenesis. ASC deletion of NR2F2, prior to adipogenesis, mimicked the reduced FAO capacity observed in ASCs from n6-FA exposed pups, suggesting that NR2F2 is required in ASCs for robust beige regulator expression and downstream nutrient oxidation in adipocytes. Transiently re-activating NR2F2 with ligand prior to differentiation in ASCs from n6-FA exposed pups, restored their FAO capacity as adipocytes by increasing the PPAR{gamma}-PGC1 axis, mitochondrial FA transporter CPT1A, ATP5 family synthases, and NDUF family Complex I proteins. Our findings suggest that excessive n6-FA exposure early in life dampens an NR2F2-mediated induction of beige adipocyte regulators, resulting in metabolic programming that is shifted towards nutrient storage.

physiology↗

Human Cytomegalovirus in breast milk is associated with milk composition, the infant gut microbiome, and infant growth

Human cytomegalovirus (CMV) is a highly prevalent herpesvirus that is often transmitted to the neonate via breast milk. Postnatal CMV transmission can have negative health consequences for preterm and immunocompromised infants, but any effects on healthy term infants are thought to be benign. Furthermore, the impact of CMV on the composition of the hundreds of bioactive factors in human milk has not been tested. Here, we utilize a cohort of exclusively breastfeeding full term mother-infant pairs to test for differences in the milk transcriptome and metabolome associated with CMV, and the impact of CMV in breast milk on the infant gut microbiome and infant growth. We find upregulation of the indoleamine 2,3-dioxygenase (IDO) tryptophan-to-kynurenine metabolic pathway in CMV+ milk samples, and that CMV+ milk is associated with decreased Bifidobacterium in the infant gut. Our data indicate a complex relationship between milk CMV, milk kynurenine, and infant growth; with kynurenine positively correlated, and CMV viral load negatively correlated, with infant weight-for-length at 1 month of age. These results suggest CMV transmission, CMV-related changes in milk composition, or both may be modulators of full term infant development.

genomics↗

Human milk variation is shaped by maternal genetics and impacts the infant gut microbiome

Human milk is a complex mix of nutritional and bioactive components that provide complete nutrition for the infant. However, we lack a systematic knowledge of the factors shaping milk composition and how milk variation influences infant health. Here, we used multi-omic profiling to characterize interactions between maternal genetics, milk gene expression, milk composition, and the infant fecal microbiome in 242 exclusively breastfeeding mother-infant pairs. We identified 487 genetic loci associated with milk gene expression unique to the lactating mammary gland, including loci that impacted breast cancer risk and human milk oligosaccharide concentration. Integrative analyses uncovered connections between milk gene expression and infant gut microbiome, including an association between the expression of inflammation-related genes with IL-6 concentration in milk and the abundance of Bifidobacteria in the infant gut. Our results show how an improved understanding of the genetics and genomics of human milk connects lactation biology with maternal and infant health.

genomics↗

Hypothalamic melanocortin-4 receptors on astrocytes mediate inflammation and body weight homeostasis

World-wide, nearly 40% of the adult population is classified as clinically obese. Central inflammation is highly correlated with obesity and increases morbidity and deterioration of health. The hypothalamus is a brain region that governs many facets of energy homeostasis, and melanocortins in the hypothalamus both decrease feeding and increase metabolism via melanocortin-4 receptors (MC4Rs). Although MC4Rs are present on neurons and astrocytes (aMC4R) previous work has focused almost exclusively on the neuronal population with the contribution of aMC4R on these processes largely unknown. Our objective was to determine the effects of hypothalamic aMC4R deletion on central and peripheral inflammation, as well as feeding and body weight homeostasis. Adult MC4R fl/fl mice were microinjected with an astrocyte-specific promoter driving Cre-expression (AAV-GFAP-GFP-Cre) or AAV-control (AAV-GFAP-GFP; n=4-7/group/sex) to produce a hypothalamic knock-down of aMC4R (KD). Body weight and composition were monitored throughout the study, and indirect calorimetry was conducted at 1 and 4 weeks after AAV injection. Acquisition of operant self-administration of palatable food was also examined. Mice were euthanized 7-8 weeks post AAV injection and brain and tissue samples were collected. We observed a significant increase in body weight, feeding, and energy balance in the KD group compared to control group. Inflammation was significantly increased centrally in KD mice within the hypothalamus, but not peripherally within serum. Additionally, aMC4R KD mice trended towards an increased reward learning for palatable food. This is the first demonstration that hypothalamic aMC4R, independent of neuronal MC4R, is important in modulating inflammation as well as contributing to energy balance. These results provide an integral understanding of the aMC4R system that will provide the foundation for future studies investigating the role of aMC4R in various disease states.

neuroscience↗