The Epigenetic Legacy of Maternal Protein Restriction: Renal Ptger1 DNA Methylation Changes in Hypertensive Rat Offspring.

Nutrient imbalances during gestation are a risk factor for hypertension in offspring. Although the effects of prenatal nutritional deficiency on the development of hypertension and cardiovascular diseases in adulthood have been extensively documented, its underlying mechanisms remain poorly understood. In this study, we aimed to elucidate the precise role and functional significance of epigenetic modifications in the pathogenesis of hypertension. To this end, we integrated methylome and transcriptome data to identify potential salt-sensitive hypertension genes using the kidneys of stroke-prone spontaneously hypertensive rat (SHRSP) pups exposed to a low-protein diet throughout their fetal life. Maternal protein restriction during gestation led to a positive correlation between DNA hypermethylation of the renal prostaglandin E receptor 1 (Ptger1) CpG island and high mRNA expression of Ptger1 in offspring, which is consistently conserved. Furthermore, post-weaning low-protein or high-protein diets modified the Ptger1 DNA hypermethylation caused by fetal malnutrition. Here, we show that this epigenetic variation in Ptger1 is linked to disease susceptibility established during fetal stages and could be reprogrammed by manipulating the postnatal diet. Thus, our findings clarify the developmental origins connecting the maternal nutritional environment and potential epigenetic biomarkers for offspring hypertension. These findings shed light on hypertension prevention and prospective therapeutic strategies.

Effects of Heat-Moisture-Treated High-Amylose Rice Flour on Body Weight, Lipid Metabolism, and Gut Microbiome Composition in Obese Rats.

The rising prevalence of lifestyle diseases, such as type 2 diabetes, cardiovascular diseases, and metabolic syndrome, has increased the need for effective dietary interventions. This study aimed to evaluate the effects of heat-moisture-treated high-amylose rice (HA-HMT) on body weight, lipid metabolism, and gut microbiome composition in a rat model of obesity. Starch digestibility-specifically, resistant starch-has been shown to provide various health benefits, including improved metabolic health and gut microbiome composition. We employed a sequential approach: firstly, utilizing diet-induced obesity rat models fed with HMT-processed and HMT-non-processed low- or high-amylose rice to investigate the potential of amylose content or HMT to alter phenotypic characteristics and lipid metabolism; and secondly, using the optimal rice flour identified in the previous step to explore the underlying mechanisms. Our findings indicate that heat-moisture treatment, rather than the level of the amylose content of the rice, contributes to the observed anti-obesity and cholesterol-lowering effects. We identified candidate genes contributing to the cholesterol-regulating potential and demonstrated that HMT rice flour could influence the gut microbiome, particularly the Ruminococcus taxa. This study provides valuable insights into the health benefits of HA-HMT rice and supports its potential as a functional food ingredient in the management of obesity and cholesterol-related disorders.

Effects of Thyme (Thymus vulgaris L.) Essential Oil on Aging-Induced Brain Inflammation and Blood Telomere Attrition in Chronologically Aged C57BL/6J Mice.

Chronological aging is commonly accompanied by chronic low-grade inflammation (or "inflammaging"), a contributor to the development of age-related chronic diseases. Aging increases oxidative stress that accelerates telomere shortening, leading to cell senescence and the generation of senescence-associated secretory phenotype (SASP) that exacerbates inflammation. Dietary antioxidants may help protect telomeres and attenuate inflammation. Thyme essential oil (TEO), reported for its potency against neuroinflammation, was fed to chronologically aged C57BL/6J mice for 24 weeks. The TEO diet showed notable impacts on the hippocampus, indicated by lower expression of the aging-related gene p16INK4A (p = 0.0783) and significantly lower expression of cyclin D kinase Cdk4 and Cdk6 (p < 0.05) compared to the age-matched control mice. The TEO group also showed significantly lower gene expression of the pro-inflammatory cytokine Il6 (p < 0.05) in the hippocampus and lower Il1b expression in the liver and cerebellum (p < 0.05). In vitro experiments conducted on NIH-3T3 cells expressing SASP revealed the dose-dependent anti-inflammatory activity of TEO. Remarkably, TEO diet-fed mice showed higher survival rates and significantly longer blood telomere lengths than the control mice. Monoterpene antioxidants in TEO, particularly thymol and p-cymene, may primarily contribute to the anti-inflammatory and telomere-protecting activities of TEO.

Postnatal nutrition environment reprograms renal DNA methylation patterns in offspring of maternal protein-restricted stroke-prone spontaneously hypertensive rats.

Maternal malnutrition hampers the offspring health by manipulating the epigenome. Recent studies indicate that the changes in DNA methylation could be reversed by afterbirth nutrition supplementation. In this study, we used DNA methylation arrays to comprehensively investigate the DNA methylation status of the renal promoter regions and the effects of postnatal protein intake on DNA methylation. We fed stroke-prone spontaneously hypertensive (SHRSP) rat dams a normal diet or a low-protein diet during pregnancy, and their 4-week-old male offspring were fed a normal diet or a high-/low-protein diet for 2 weeks. We found that the methylation status of 2,395 differentially methylated DNA regions was reprogrammed, and 34 genes were reset by different levels of postnatal protein intake in the offspring. Among these genes, Adora2b, Trpc5, Ar, Xrcc2, and Atp1b1 are involved in renal disease and blood pressure regulation. Our findings indicate that postnatal nutritional interventions can potentially reprogram epigenetic changes, providing novel therapeutic and preventive epigenetic targets for salt-sensitive hypertension.

Effects of Turmeric Extract on Age-Related Skeletal Muscle Atrophy in Senescence-Accelerated Mice.

Muscle atrophy is one of the main causes of sarcopenia-the age-related loss of skeletal muscle. In this study, we investigated the effect of turmeric (Curcuma longa) extract (TE) supplementation on age-related muscle atrophy in a senescence-accelerated mouse model and explored the underlying mechanisms. Twenty-six-week-old male, senescence-accelerated mouse resistant (SAMR) mice received the AIN-93G basal diet, while twenty-six-week-old male, senescence-accelerated mouse prone 8 (SAMP8) mice received the AIN-93G basal diet or a 2% TE powder-supplemented diet for ten weeks. Our findings revealed that TE supplementation showed certain effects on ameliorating the decrease in body weight, tibialis anterior weight, and mesenteric fat tissue weight in SAMP8 mice. TE improved gene expression in the glucocorticoid receptor-FoxO signaling pathway in skeletal muscle, including redd1, klf15, foxo1, murf1, and mafbx. Furthermore, TE might have the certain potential on improving the dynamic balance between anabolic and catabolic processes by inhibiting the binding of glucocorticoid receptor or FoxO1 to the glucocorticoid response element or FoxO-binding element in the MuRF1 promoter in skeletal muscle, thereby promoting muscle mass and strength, and preventing muscle atrophy and sarcopenia prevention. Moreover, TE may have reduced mitochondrial damage and maintained cell growth and division by downregulating the mRNA expression of the genes mfn2 and tsc2. Thus, the results indicated TE's potential for preventing age-related muscle atrophy and sarcopenia.

Association between Dietary Behaviors and BMI Stratified by Sex and the ALDH2 rs671 Polymorphism in Japanese Adults.

The rs671 polymorphism, unique to East Asians, is well known to change the sensitivity to alcohol. Moreover, this polymorphism is associated not only with alcohol intake but also with several dietary behaviors (DBs), chronic diseases, and BMI, but the triadic association among the rs671 genotype, DBs, and BMI is unclear. This study included 12,271 Japanese subjects and aimed to observe this three-way association using the rs671 polymorphism, data of 56 DBs, and BMI. All analyses were stratified by participant sex. First, linear regression analyses resulted in significant associations between 18 and 21 DBs and BMI in males and females, respectively. Next, genetic heterogeneity was observed in all sub-groups via interaction analysis of the rs671 genotype stratified by drinking habits. Finally, we observed the characteristics of BMI-related DBs based on the rs671 genotype via stepwise regression analyses stratified by the rs671 genotype and drinking habits. Notably, positive associations were observed between lactobacillus beverage intake and BMI among participants with the rs671 polymorphism AA genotype in both sexes. This study suggests that the rs671 polymorphism modifies the association between DBs and BMI independently of drinking habits, providing evidence for the potential use of rs671 polymorphism information for precision nutrition with East Asians.

Transcriptome and Methylome Profiling in Rat Skeletal Muscle: Impact of Post-Weaning Protein Restriction.

Skeletal muscle is programmable, and early-life nutritional stimuli may form epigenetic memory in the skeletal muscle, thus impacting adult muscle function, aging, and longevity. In the present study, we designed a one-month protein restriction model using post-weaning rats, followed by a two-month rebound feeding, to investigate how early-life protein restriction affects overall body growth and muscle development and whether these influences could be corrected by rebound feeding. We observed comprehensive alterations immediately after protein restriction, including retarded growth, altered biochemical indices, and disturbed hormone secretion. Transcriptome profiling of the gastrocnemius muscle followed by gene ontology analyses revealed that "myogenic differentiation functions" were upregulated, while "protein catabolism" was downregulated as a compensatory mechanism, with enhanced endoplasmic reticulum stress and undesired apoptosis. Furthermore, methylome profiling of the gastrocnemius muscle showed that protein restriction altered the methylation of apoptotic and hormone secretion-related genes. Although most of the alterations were reversed after rebound feeding, 17 genes, most of which play roles during muscle development, remained altered at the transcriptional level. In summary, early-life protein restriction may undermine muscle function in the long term and affect skeletal muscle development at the both transcriptional and methylation levels, which may hazard future muscle health.

Suppressive Effects of Turmeric Extract on Muscle Atrophy in Dexamethasone-Treated Mice and Myotubes.

Sarcopenia is the decline in skeletal muscle mass, strength, and functions, which decreases the quality of life in elderly people. This study investigated the suppressive effect of turmeric (Curcuma longa) extract (TE) on muscle atrophy in dexamethasone (DEX)-treated mice and C2C12 myotubes. DEX treatment significantly decreased the muscle weight and significantly increased Fbxo32 and Murf1 expression in mice, and these changes were suppressed by the supplementation of an AIN-93 based diet with 2% TE. A similar pattern was observed in FBXO32 and MuRF1 protein expression. In C2C12 myotubes, DEX treatment significantly increased FBXO32 and MuRF1 gene and protein expression, and these increases were significantly suppressed by TE supplementation at a concentration of 200 µg/mL. Furthermore, one of the five TE fractions, which were separated by high-performance liquid chromatography had a similar effect with TE supplementation. The present study proposes the suppressive effect of turmeric on sarcopenia.

Effect of heat-moisture treated brown rice crackers on postprandial flow-mediated dilation in adults with mild endothelial dysfunction.

Background: Endothelial dysfunction is an early pathophysiological feature and independent predictor of a poor prognosis in most forms of cardiovascular disease. We evaluated the effect of brown rice crackers (BR-C) on endothelial function. Methods: Effect of heat-moisture treated (HMT) -BR-C on postprandial flow-mediated dilation (FMD) in adults with mild endothelial dysfunction was compared with that of BR-C and white rice crackers (WR-C) in 12 adults with mild endothelial dysfunction (less than 7.0% of FMD) by a randomized, single-blind, three-treatment three-period crossover trial (UMIN 000034898). Since we considered that the FMD increase was associated with the treatment of HMT-BR-C, we examined the effect of three possible factors: postprandial glucose levels, polyphenol content, and polyphenol release from the food matrix. Results: Mean pre-intake baseline FMD values of HMT-BR-C, BR-C, and WR-C were 4.9%, 5.1%, and 4.9%, respectively, and those values 1 h post-intake were 6.3%, 5.1%, and 4.8%, respectively. There was no difference in intergroup comparisons of FMD using Dunnett's multiple comparison test. There was a significant increase in FMD only in HMT-BR-C in intragroup comparisons (P = 0.042 by paired-t test). In comparison with BR-C, no significant difference was noted in the postprandial glucose level nor in the content of total polyphenols and ferulic acid derivatives in HMT-BR-C. However, the 70% ethanol extracted from HMT-BR-C contained a significantly larger amount of free and bound ferulic acids than from BR-C. Conclusion: HMT-BR-C intake increased the postprandial FMD response.

Activating transcription factor 4-dependent hsa-miR-663a transcription mediates mTORC1/p70S6K1 signaling underleucine deprivation.

The mechanistic target of rapamycin complex 1 (mTORC1) is involved in nutrient-induced signaling and is a master regulator of cell growth and metabolism. Amino acid-deficient conditions affect mTORC1 activity; however, its upstream regulators warrant further investigation. MicroRNAs are key regulators of nutrient-related responses; therefore, the present study aimed to assess the leucine starvation-induced microRNA profile and its impact on mTORC1 activity. Transcriptome analysis of human hepatocellular carcinoma cells (HepG2) under leucine deprivation revealed that hsa-miR-663a and hsa-miR-1469 were altered in a transcription factor 4-dependent manner. Overexpression of these microRNAs induced phosphorylation of the ribosomal protein S6 kinase beta-1, a mTORC1 downstream target. Furthermore, hsa-miR-663a downregulated proline-rich Akt1 substrate of 40 kDa (PRAS40), one of the mTORC1 components. In summary, this study provides new insights into the regulatory role of microRNAs in amino acid metabolism and demonstrates alterations in microRNA profile under leucine deprivation in human hepatocytes.

Dietary Eggshell Membrane Powder Improves Survival Rate and Ameliorates Gut Dysbiosis in Interleukin-10 Knockout Mice.

Inflammatory bowel disease (IBD) is known to be associated with compositional and metabolic changes in the gut microbiota. The aim of this study was to investigate whether dietary eggshell membrane (ESM) improves survival rate or ameliorates gut dysbiosis in a spontaneous IBD model of interleukin-10 knockout (IL10-/-) mice. Female C57BL/6J wild-type (WT) and IL10-/- mice (KO) were fed an AIN-93G basal diet or an ESM diet (KOE) for 19 weeks. Gut microbiota profiles were analyzed via 16S rRNA sequencing, and short-chain fatty acids in cecal content were analyzed with high-performance liquid chromatography. The results demonstrated that ESM supplementation significantly improved the survival rate and body composition in KO mice. Alpha diversity analysis of the microbiota revealed that ESM supplementation significantly increased gut microbial diversity, which was decreased in IL10-/- mice. The Firmicutes/Bacteroidetes ratio was recovered to a normal level by ESM supplementation, suggesting that ESM helps maintain the compositional balance of the gut microbiota. ESM increased relative abundance of commensal bacterial Ruminococcus and Bacteroidales S24-7 and reduced the abundance of the proinflammatory-related bacterium, Enterobacteriaceae. Additionally, ESM supplementation promoted the production of butyrate in cecal contents and downregulated the expression of proinflammatory genes, including interleukin-1ß (Il-1ß) and tumor necrosis factor-α (Tnf-α) in IL10-/- mice colon, indicating anti-inflammatory functions. These findings suggest that ESM may be used as a beneficial dietary intervention for IBD.

Eggshell membrane modulates gut microbiota to prevent murine pre-cachexia through suppression of T helper cell differentiation.

BACKGROUND: Cachexia is a life-threatening condition observed in several pathologies, such as cancer or chronic diseases. Interleukin 10 (Il10) gene transfer is known to improve cachexia by downregulating Il6. Here, we used an IL10-knockout mouse model to simulate cachexia and investigate the effects of eggshell membrane (ESM), a resistant protein, on general pre-cachexia symptoms, which is particularly important for the development of cachexia therapeutics. METHODS: Five-week-old male C57BL6/J mice were fed an AIN-93G powdered diet (WT), and 5-week-old male B6.129P2-Il10 < tm1Cgn>/J (IL10-/- ) mice were fed either the AIN-93G diet (KO) or an 8% ESM-containing diet (KOE) for 28 weeks. The tissue weight and levels of anaemia-, blood glucose-, lipid metabolism-, and muscular and colonic inflammation-related biochemical markers were measured. Transcriptomic analysis on liver and colon mucus and proteomic analysis on skeletal muscle were performed. Ingenuity Pathway Analysis was used to identify molecular pathways and networks. Caecal short-chain fatty acids (SCFAs) were identified using HPLC, and caecal bacteria DNA were subjected to metagenomic analysis. Flow cytometry analysis was performed to measure the CD4+ IL17+ T cells in mesenteric lymph nodes. RESULTS: The body weight, weight of gastrocnemius muscle and fat tissues, colon weight/length ratio, plasma HDL and NEFA, muscular PECAM-1 levels (P < 0.01), plasma glucose and colonic mucosal myeloperoxidase activity (P < 0.05) and T helper (Th) 17 cell abundance (P = 0.071) were improved in KOE mice over KO mice. Proteomic analysis indicated the protective role of ESM in muscle weakness and maintenance of muscle formation (>1.5-fold). Transcriptomic analysis revealed that ESM supplementation suppressed the LPS/IL1-mediated inhibition of RXR function pathway in the liver and downregulated the colonic mucosal expression of chemokines and Th cell differentiation-related markers (P < 0.01) by suppressing the upstream BATF pathway. Analysis of the intestinal microenvironment revealed that ESM supplementation ameliorated the microbial alpha diversity and the abundance of microbiota associated with the degree of inflammation (P < 0.05) and increased the level of total organic acids, particularly of SCFAs such as butyrate (2.3-fold), which could inhibit Th1 and Th17 production. CONCLUSIONS: ESM supplementation ameliorated the chief symptoms of cachexia, including anorexia, lean fat tissue mass, skeletal muscle wasting and reduced physical function. ESM also improved colon and skeletal muscle inflammation, lipid metabolism and microbial dysbiosis. These results along with the suppressed differentiation of Th cells could be associated with the beneficial intestinal microenvironment and, subsequently, attenuation of pre-cachexia. Our findings provide insights into the potential of ESM in complementary interventions for pre-cachexia prevention.

Laurel Attenuates Dexamethasone-Induced Skeletal Muscle Atrophy In Vitro and in a Rat Model.

Prevention of muscle atrophy contributes to improved quality of life and life expectancy. In this study, we investigated the effects of laurel, selected from 34 spices and herbs, on dexamethasone (DEX)-induced skeletal muscle atrophy and deciphered the underlying mechanisms. Co-treatment of C2C12 myotubes with laurel for 12 h inhibited the DEX-induced expression of intracellular ubiquitin ligases-muscle atrophy F-box (atrogin-1/MAFbx) and muscle RING finger 1 (MuRF1)-and reduction in myotube diameter. Male Wistar rats were supplemented with 2% laurel for 17 days, with DEX-induced skeletal muscle atrophy occurring in the last 3 days. Laurel supplementation inhibited the mRNA expression of MuRF1, regulated DNA damage and development 1 (Redd1), and forkhead box class O 1 (Foxo1) in the muscles of rats. Mechanistically, we evaluated the effects of laurel on the cellular proteolysis machinery-namely, the ubiquitin/proteasome system and autophagy-and the mTOR signaling pathway, which regulates protein synthesis. These data indicated that the amelioration of DEX-induced skeletal muscle atrophy induced by laurel, is mainly mediated by the transcriptional inhibition of downstream factors of the ubiquitin-proteasome system. Thus, laurel may be a potential food ingredient that prevents muscle atrophy.

iTRAQ-Based Quantitative Proteomics Reveals the Energy Metabolism Alterations Induced by Chlorogenic Acid in HepG2 Cells.

Epidemiological studies have suggested that coffee consumption is associated with a decrease in the risk of developing obesity and diabetes; however, the detailed mechanisms underlying these effects of coffee consumption remain poorly understood. In this study, we examined the effects of chlorogenic acid on energy metabolism in vitro. Hepatocellular carcinoma G2 (HepG2) cells were cultured in a medium containing chlorogenic acid. Chlorogenic acid increased the activity of mitochondrial enzymes, including citrate synthase, isocitrate dehydrogenase, and malate dehydrogenase (MDH), which are involved in the tricarboxylic acid (TCA) cycle. Proteome analysis using the isobaric tags for the relative and absolute quantitation (iTRAQ) method revealed the upregulation of proteins involved in the glycolytic system, electron transport system, and ATP synthesis in mitochondria. Therefore, we propose a notable mechanism whereby chlorogenic acid enhances energy metabolism, including the TCA cycle, glycolytic system, electron transport, and ATP synthesis. This mechanism provides important insights into understanding the beneficial effects of coffee consumption.

Maternal Protein Restriction and Post-Weaning High-Fat Feeding Alter Plasma Amino Acid Profiles and Hepatic Gene Expression in Mice Offspring.

Maternal undernutrition during pregnancy is closely associated with epigenetic changes in the child, and it affects the development of obesity throughout the child's life. Here, we investigate the effect of fetal low protein exposure and post-weaning high-fat consumption on plasma amino acid profiles and hepatic gene expression. Mother C57BL/6J mice were fed a 20% (CN) or 9% (LP) casein diet during pregnancy. After birth, the male offspring of both these groups were fed a high-fat diet (HF) from 6 to 32 weeks. At 32 weeks, the final body weight between the two groups remained unchanged, but the LP-HF group showed markedly higher white fat weight and plasma leptin levels. The LP-HF group exhibited a significant increase in the concentrations of isoleucine, leucine, histidine, phenylalanine, serine, and tyrosine. However, no differences were observed in the lipid content in the liver. According to the hepatic gene expression analysis, the LP-HF group significantly upregulated genes involved in the chromatin modification/organization pathways. Thus, maternal low protein and a post-weaning high-fat environment contributed to severe obesity states and changes in gene expression related to hepatic chromatin modification in offspring. These findings provide novel insights for the prevention of lifestyle-related diseases at the early life stage.

The Potential Roles of Probiotics, Resistant Starch, and Resistant Proteins in Ameliorating Inflammation during Aging (Inflammaging).

Aging is typically accompanied by biological and physiological changes that alter cellular functions. Two of the most predominant phenomena in aging include chronic low-grade inflammation (inflammaging) and changes in the gut microbiota composition (dysbiosis). Although a direct causal relationship has not been established, many studies have reported significant reductions in inflammation during aging through well-maintained gut health and microbial balance. Prebiotics and probiotics are known to support gut health and can be easily incorporated into the daily diet. Unfortunately, few studies specifically focus on their significance in reducing inflammation during aging. Therefore, this review summarizes the scientific evidence of the potential roles of probiotics and two types of prebiotics, resistant starch and resistant proteins, in later age. Studies have demonstrated that the oral consumption of bacteria that may contribute to anti-inflammatory response, such as Bifidobacterium spp., Akkermansia munichipilla, and Faecalis praunitzii, contributes significantly to the suppression of pro-inflammatory markers in elderly humans and aged animals. Colonic fermentation of resistant starch and proteins also demonstrates anti-inflammatory activity owing to the production of butyrate and an improvement in the gut microbiota composition. Collectively, probiotics, resistant starch, and resistant proteins have the potential to promote healthy aging.

Ameliorating Effects of Coriander on Gastrocnemius Muscles Undergoing Precachexia in a Rat Model of Rheumatoid Arthritis: A Proteomics Analysis.

Coriander is a commonly used vegetable, spice, and folk medicine, possessing both nutritional and medicinal properties. Up to two-thirds of patients with rheumatoid arthritis (RA) exhibit loss of body mass, predominately skeletal muscle mass, a process called rheumatoid cachexia, and this has major effects of the quality of life of patients. Owing to a lack of effective treatments, the initial stage of cachexia has been proposed as an important period for prevention and decreasing pathogenesis. In the current study, we found that cachexia-like molecular disorders and muscle weight loss were in progress in gastrocnemius muscle after only 5 days of RA induction in rats, although rheumatoid cachexia symptoms have been reported occurring approximately 45 days after RA induction. Oral administration of coriander slightly restored muscle loss. Moreover, iTRAQ-based quantitative proteomics revealed that coriander treatment could partially restore the molecular derangements induced by RA, including impaired carbon metabolism, deteriorated mitochondrial function (tricarboxylic acid cycle and oxidative phosphorylation), and myofiber-type alterations. Therefore, coriander could be a promising functional food and/or complementary therapy for patients with RA against cachexia.

Effects of Eggshell Membrane on Keratinocyte Differentiation and Skin Aging In Vitro and In Vivo.

Skin aging is one of the hallmarks of the aging process that causes physiological and morphological changes. Recently, several nutritional studies were conducted to delay or suppress the aging process. This study investigated whether nutritional supplementation of the eggshell membrane (ESM) has a beneficial effect on maintaining skin health and improving the skin aging process in vitro using neonatal normal human epidermal keratinocytes (NHEK-Neo) and in vivo using interleukin-10 knockout (IL-10 KO) mice. In NHEK-Neo cells, 1 mg/mL of enzymatically hydrolyzed ESM (eESM) upregulated the expression of keratinocyte differentiation markers, including keratin 1, filaggrin and involucrin, and changed the keratinocyte morphology. In IL-10 KO mice, oral supplementation of 8% powdered-ESM (pESM) upregulated the expression of growth factors, including transforming growth factor ß1, platelet-derived growth factor-ß and connective tissue growth factor, and suppressed skin thinning. Furthermore, voltage-gated calcium channel, transient receptor potential cation channel subfamily V members were upregulated by eESM treatment in NHEK-Neo cells and pESM supplementation in IL-10 KO mice. Collectively, these data suggest that ESM has an important role in improving skin health and aging, possibly via upregulating calcium signaling.