Mulberry (Morus alba L.) leaf is a well-established traditional Chinese botanical and culinary resource. It has found widespread application in the management of diabetes. The bioactive constituents of mulberry leaf, specifically mulberry leaf flavonoids (MLFs), exhibit pronounced potential in the amelioration of type 2 diabetes (T2D). This potential is attributed to their ability to safeguard pancreatic β cells, enhance insulin resistance, and inhibit α-glucosidase activity. Our antecedent research findings underscore the substantial therapeutic efficacy of MLFs in treating T2D. However, the precise mechanistic underpinnings of MLF's anti-T2D effects remain the subject of inquiry. Activation of brown/beige adipocytes is a novel and promising strategy for T2D treatment. In the present study, our primary objective was to elucidate the impact of MLFs on adipose tissue browning in db/db mice and 3T3-L1 cells and elucidate its underlying mechanism. The results manifested that MLFs reduced body weight and food intake, alleviated hepatic steatosis, improved insulin sensitivity, and increased lipolysis and thermogenesis in db/db mice. Moreover, MLFs activated brown adipose tissue (BAT) and induced the browning of inguinal white adipose tissue (IWAT) and 3T3-L1 adipocytes by increasing the expressions of brown adipocyte marker genes and proteins such as uncoupling protein 1 (UCP1) and beige adipocyte marker genes such as transmembrane protein 26 (Tmem26), thereby promoting mitochondrial biogenesis. Mechanistically, MLFs facilitated the activation of BAT and the induction of WAT browning to ameliorate T2D primarily through the activation of AMP-activated protein kinase (AMPK)/sirtuin 1 (SIRT1)/peroxisome proliferator-activated receptor-gamma coactivator 1α (PGC-1α) signaling pathway. These findings highlight the unique capacity of MLF to counteract T2D by enhancing BAT activation and inducing browning of IWAT, thereby ameliorating glucose and lipid metabolism disorders. As such, MLFs emerge as a prospective and innovative browning agent for the treatment of T2D.
Mice ; Animals ; Adipose Tissue, Brown ; Sirtuin 1/pharmacology* ; Diabetes Mellitus, Type 2/metabolism* ; AMP-Activated Protein Kinases/metabolism* ; Morus/metabolism* ; Flavonoids/metabolism* ; Prospective Studies ; Signal Transduction ; Adipose Tissue, White ; Plant Leaves ; Uncoupling Protein 1/metabolism* ; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha/metabolism*

This study investigated the mechanism of Zexie Decoction(ZXD) in promoting white adipose tissue browning/brown adipose tissue activation based on the GLP-1R/cAMP/PKA/CREB pathway. A hyperlipidemia model was induced by a western diet(WD) in mice, and the mice were divided into a control group, a model group(WD), and low-, medium-, and high-dose ZXD groups. An adipogenesis model was induced in 3T3-L1 cells in vitro, and with forskolin(FSK) used as a positive control, low-, medium-, and high-dose ZXD groups were set up. Immunohistochemistry and immunofluorescence results showed that compared with the WD group, ZXD promoted the expression of UCP1 in white and brown adipose tissues, and also upregulated UCP1, CPT1β, PPARα, and other genes in the cells. Western blot analysis showed a dose-dependent increase in the protein expression of PGC-1α, UCP1, and PPARα with ZXD treatment, indicating that ZXD could promote the white adipose tissue browning/brown adipose tissue activation. Hematoxylin-eosin(HE) staining results showed that after ZXD treatment, white and brown adipocytes were significantly reduced in size, and the mRNA expression of ATGL, HSL, MGL, and PLIN1 was significantly upregulated as compared with the results in the WD group. Oil red O staining and biochemical assays indicated that ZXD improved lipid accumulation and promoted lipolysis. Immunohistochemistry and immunofluorescence staining for p-CREB revealed that ZXD reversed the decreased expression of p-CREB caused by WD. In vitro intervention with ZXD increased the protein expression of CREB, p-CREB, and p-PKA substrate, and increased the mRNA level of CREB. ELISA detected an increase in intracellular cAMP concentration with ZXD treatment. Molecular docking analysis showed that multiple active components in Alismatis Rhizoma and Atractylodis Macrocephalae Rhizoma could form stable hydrogen bond interactions with GLP-1R. In conclusion, ZXD promotes white adipose tissue browning/brown adipose tissue activation both in vivo and in vitro, and its mechanism of action may be related to the GLP-1R/cAMP/PKA/CREB pathway.
Mice ; Animals ; Adipose Tissue, Brown ; Molecular Docking Simulation ; PPAR alpha/metabolism* ; Adipose Tissue, White ; RNA, Messenger/metabolism*

adipose tissue (BAT) from white adipose tissue (WAT) in rats using spectral computed tomography (CT) with histological validation.MATERIALS AND METHODS: A lipid-containing phantom (lipid fractions from 0% to 100%) was imaged with spectral CT. An in vivo, non-enhanced spectral CT scan was performed on 24 rats, and fat concentrations of BAT and WAT were measured. The rats were randomized to receive intraperitoneal treatment with norepinephrine (NE) (n = 12) or saline (n = 12). Non-enhanced and enhanced spectral CT scans were performed after treatment to measure the elevation of iodine in BAT and WAT. The BAT/aorta and WAT/aorta ratios were calculated and compared, after which isolated BAT and WAT samples were subjected to histological and uncoupling protein 1 (UCP1) analyses.RESULTS: The ex-vivo phantom study showed excellent linear fit between measured fat concentration and the known gravimetric reference standard (r² = 0.996). In vivo, BAT had significantly lower fat concentration than WAT (p < 0.001). Compared to the saline group, the iodine concentration of BAT increased significantly (p < 0.001) after injection of NE, while the iodine concentration of WAT only changed slightly. The BAT/aorta ratio also increased significantly after exposure to NE compared to the saline group (p < 0.001). Histological and UCP1 expression analyses supported the spectral CT imaging results.CONCLUSION: The study consolidates spectral CT as a new approach for non-invasive imaging of BAT and WAT. Quantitative analyses of BAT and WAT by spectral CT revealed different characteristics and pharmacologic activations in the two types of adipose tissue.]]>
Adipose Tissue ; Adipose Tissue, Brown ; Adipose Tissue, White ; Animals ; Iodine ; Norepinephrine ; Rats ; Tomography, X-Ray Computed

To characterize the timeliness of β3 adrenergic receptor agonist CL316,243-induced browning of white adipose tissues in mice.
 Methods: Male C57BL/6J mice at 10 weeks of age were housed in conventional cages and given sterile saline for the control group or CL316,243 (1 μg/g) for the experimental group via intraperitoneal injection for 1, 3, and 5 days. Food intake and body weight were measured daily. Interscapular brown adipose tissue (iBAT), inguinal subcutaneous white adipose (sWAT) and epididymal white adipose tissue (eWAT) were harvested for histological and gene expression analysis.
 Results: Compared with the control group, intraperitoneal injection of CL316,243 reduced the weight of eWAT on the first day. Meanwhile, CL316,243 continuously promoted the mRNA and protein expression of uncoupling protein-1 (UCP-1) in sWAT and eWAT. Furthermore, CL316,243 injection significantly decreased the food intake and weight gain of the mice, and reduced the diameter of adipocyte and accumulation of small lipid droplets in adipose tissues.
 Conclusion: CL316,243 can induce the brown-like remodeling in adipose tissues of mice in vivo, which show different time-dependent manners in different adipose tissues.
Adipose Tissue, Brown ; Adipose Tissue, White ; Adrenergic beta-Agonists ; Animals ; Male ; Mice ; Mice, Inbred C57BL ; Uncoupling Protein 1

OBJECTIVE: To investigate the time-sequential expression of a novel long non-coding RNA, lnc AK079912, in metabolically related tissues and during adipose tissue development and browning in mice. METHODS: The interscapular brown adipose tissue (iBAT), subcutaneous white adipose tissue (sWAT), epididymal white adipose tissue (eWAT), liver tissues and muscular tissues were collected from 8-week-old C57BL/6J mice. The iBAT, sWAT and eWAT were also collected from the mice during development (0 day, 21 days, 8 weeks and 6 months after birth) and from 8- to 10-week- mice with cold exposure (4 ℃) and intraperitoneal injections of CL316, 243 (1 μg/g body weight) for 1 to 5 days. Trizol was used to extract the total RNA from the tissues, and RT-qPCR was performed to detect the expressions of lnc AK079912. Isolated mouse preadipocytes in primary culture were induced for adipogenic differentiation for 9 days and then treated with CL316, 243 (2 μmol/L) for different durations (no longer than 24 h); the expression of lnc AK079912 in the cells was detected using RT-qPCR at different time points of the treatment. RESULTS: Lnc AK079912 was highly expressed in mouse adipose tissues, the highest in iBAT, followed by the muscular tissue, but was hardly detected in the liver tissue. The expression level of lnc AK079912 increased progressively in iBAT and sWAT during development of the mice, while its expression in eWAT showed an initial increase followed by a reduction at 8 weeks ( < 0.001). No significant difference was found in the expression of lnc AK079912 in the iBAT, sWAT or eWAT in mice with cold stimulation for 1 to 5 days ( > 0.05). The expression of lnc AK079912 was significantly decreased in iBAT and eWAT ( < 0.05) but increased in eWAT from mice with intraperitoneal injection of CL316, 243 for 1 to 5 days ( < 0.05). The expression level in the adipocytes in primary culture was significantly increased in response to treatment with CL316, 243 ( < 0.05). CONCLUSIONS Lnc AK079912 is highly expressed in mouse adipose tissue, and its expression gradually increases with the development of adipose tissue but with a depot-specific difference. Lnc AK079912 is significantly elevated in the early stage of adipose tissue browning, indicating its important role in the development and browning of adipose tissue.
Adipocytes ; Adipogenesis ; Adipose Tissue, Brown ; Adipose Tissue, White ; Animals ; Male ; Mice ; Mice, Inbred C57BL ; RNA, Long Noncoding

OBJECTIVE: We aimed to explore how fermented barley extracts with Lactobacillus plantarum dy-1 (LFBE) affected the browning in adipocytes and obese rats. METHODS: In vitro, 3T3-L1 cells were induced by LFBE, raw barley extraction (RBE) and polyphenol compounds (PC) from LFBE to evaluate the adipocyte differentiation. In vivo, obese SD rats induced by high fat diet (HFD) were randomly divided into three groups treated with oral gavage: (a) normal control diet with distilled water, (b) HFD with distilled water, (c) HFD with 800 mg LFBE/kg body weight (bw). RESULTS: In vitro, LFBE and the PC in the extraction significantly inhibited adipogenesis and potentiated browning of 3T3-L1 preadipocytes, rather than RBE. In vivo, we observed remarkable decreases in the body weight, serum lipid levels, white adipose tissue (WAT) weights and cell sizes of brown adipose tissues (BAT) in the LFBE group after 10 weeks. LFBE group could gain more mass of interscapular BAT (IBAT) and promote the dehydrogenase activity in the mitochondria. And LFBE may potentiate process of the IBAT thermogenesis and epididymis adipose tissue (EAT) browning via activating the uncoupling protein 1 (UCP1)-dependent mechanism to suppress the obesity. CONCLUSION These results demonstrated that LFBE decreased obesity partly by increasing the BAT mass and the energy expenditure by activating BAT thermogenesis and WAT browning in a UCP1-dependent mechanism.
3T3 Cells ; Adipocytes ; drug effects ; physiology ; Adipose Tissue, Brown ; drug effects ; physiology ; Adipose Tissue, White ; drug effects ; physiology ; Animal Feed ; analysis ; Animals ; Anti-Obesity Agents ; administration & dosage ; metabolism ; Cell Differentiation ; drug effects ; Diet ; Fermentation ; Hordeum ; chemistry ; Lactobacillus plantarum ; chemistry ; Male ; Mice ; Obesity ; drug therapy ; genetics ; Plant Extracts ; chemistry ; Probiotics ; administration & dosage ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Uncoupling Protein 1 ; genetics ; metabolism

OBJECTIVE: To investigate the effects of genipin on promoting brown adipose tissue activation and white adipose tissue browning. METHODS: The male C57BL/6J mice were divided into three groups: normal control group, genipin group and cold-stimulus group.Genipin group were treated consecutively with genipin at a dose of 15 mg/kg once a day for 9 days, normal control group were treated with the saline.The mice with cold-stimulus were exposed to 4℃ environment for 5 days.Daily food amount and body weight were measured.Morphological changes were observed in the subscapular region, inguinal region and epididymis around the adipose tissue.The expression of uncoupling protein 1 (UCP1) was determined by real-time PCR and Western blot respectively. RESULTS: The wet weight of white fat in genipin-treated mice was decreased by 16% , and 28% in that of cold-stimulus mice, compared with the normal control group (P＜0.05).After treatments of genipin and cold-stimulus, the color of white adipose tissues was darker, and the size of lipid droplets in adipocytes was smaller, whereas the number was increased.Compared with the normal control group, UCP1 expression was increased obviously in fat tissues, including the subcutaneous and visceral white adipose tissues, and brown adipose tissue after treated with genipin and cold-stimulus (P＜0.05). CONCLUSION Genipin promoted activation of brown adipose tissue and browning of white adipose tissue by upregulating UCP1 expression, which could contribute to the loss of body weight against obesity.
Adipose Tissue, Brown ; drug effects ; Adipose Tissue, White ; drug effects ; Animals ; Cholagogues and Choleretics ; pharmacology ; Iridoids ; pharmacology ; Male ; Mice ; Mice, Inbred C57BL ; Obesity ; drug therapy ; Uncoupling Protein 1 ; drug effects ; Up-Regulation

Adipose tissue inflammation is considered a major contributing factor in the development of obesity-associated insulin resistance and cardiovascular diseases. However, the cause of adipose tissue inflammation is presently unclear. The role of mitochondria in white adipocytes has long been neglected because of their low abundance. However, recent evidence suggests that mitochondria are essential for maintaining metabolic homeostasis in white adipocytes. In a series of recent studies, we found that mitochondrial function in white adipocytes is essential to the synthesis of adiponectin, which is the most abundant adipokine synthesized from adipocytes, with many favorable effects on metabolism, including improvement of insulin sensitivity and reduction of atherosclerotic processes and systemic inflammation. From these results, we propose a new hypothesis that mitochondrial dysfunction in adipocytes is a primary cause of adipose tissue inflammation and compared this hypothesis with a prevailing concept that “adipose tissue hypoxia” may underlie adipose tissue dysfunction in obesity. Recent studies have emphasized the role of the mitochondrial quality control mechanism in maintaining mitochondrial function. Future studies are warranted to test whether an inadequate mitochondrial quality control mechanism is responsible for mitochondrial dysfunction in adipocytes and adipose tissue inflammation.
11-beta-Hydroxysteroid Dehydrogenases ; Adipocytes ; Adipocytes, White ; Adipokines ; Adiponectin ; Adipose Tissue ; Anoxia ; Cardiovascular Diseases ; Homeostasis ; Inflammation ; Insulin Resistance ; Metabolism ; Mitochondria ; Nitric Oxide ; Obesity ; Quality Control

The global obesity epidemic and associated metabolic diseases require alternative biological targets for new therapeutic strategies. In this study, we show that a phytochemical sulfuretin suppressed adipocyte differentiation of preadipocytes and administration of sulfuretin to high fat diet-fed obese mice prevented obesity and increased insulin sensitivity. These effects were associated with a suppressed expression of inflammatory markers, induced expression of adiponectin, and increased levels of phosphorylated ERK and AKT. To elucidate the molecular mechanism of sulfuretin in adipocytes, we performed microarray analysis and identified activating transcription factor 3 (Atf3) as a sulfuretin-responsive gene. Sulfuretin elevated Atf3 mRNA and protein levels in white adipose tissue and adipocytes. Consistently, deficiency of Atf3 promoted lipid accumulation and the expression of adipocyte markers. Sulfuretin’s but not resveratrol’s anti-adipogenic effects were diminished in Atf3 deficient cells, indicating that Atf3 is an essential factor in the effects of sulfuretin. These results highlight the usefulness of sulfuretin as a new anti-obesity intervention for the prevention of obesity and its associated metabolic diseases.
Activating Transcription Factor 3 ; Adipocytes ; Adiponectin ; Adipose Tissue, White ; Animals ; Diet* ; Insulin Resistance ; Metabolic Diseases* ; Mice ; Mice, Obese* ; Microarray Analysis ; Obesity* ; RNA, Messenger

BACKGROUND/OBJECTIVES: Fasting and postprandial hyperglycemia should be controlled to avoid complications of diabetes mellitus. This study investigated the effects of autumn olive (Elaeagnus umbellata Thunb.) berry (AOB) on fasting and postprandial hyperglycemia in mice. MATERIALS/METHODS: In vitro α-glucosidase inhibitory effect of AOB was determined. Maltose solution (2 g/kg) with and without AOB extract at 500 mg/kg or acarbose at 50 mg/kg was orally administered to normal mice after overnight fasting and glucose levels were measured. To study the effects of chronic consumption of AOB, db/db mice received the basal diet or a diet containing AOB extract at 0.4% or 0.8%, or acarbose at 0.04% for 7 weeks. Blood glycated hemoglobin and serum glucose and insulin levels were measured. Expression of adiponectin protein in epididymal white adipose tissue was determined by Western blotting. RESULTS: In vitro inhibitory effect of AOB extract on α-glucosidase was 92% as strong as that of acarbose. The AOB extract (500 mg/kg) or acarbose (50 mg/kg) significantly suppressed the postprandial rise of blood glucose after maltose challenge and the area under the glycemic response curve in normal mice. The AOB extract at 0.4% or 0.8% of diet or acarbose at 0.04% of diet significantly lowered levels of serum glucose and blood glycated hemoglobin and homeostasis model assessment for insulin resistance values in db/db mice. The expression of adiponectin protein in adipose tissue was significantly elevated by the consumption of AOB at 0.8% of diet. CONCLUSIONS: Autumn olive (E. umbellata Thunb.) berry may reduce postprandial hyperglycemia by inhibiting α-glucosidase in normal mice. Chronic consumption of AOB may alleviate fasting hyperglycemia in db/db mice partly by inhibiting α-glucosidase and upregulating adiponectin expression.
Acarbose ; Adiponectin ; Adipose Tissue ; Adipose Tissue, White ; Animals ; Blood Glucose ; Blotting, Western ; Diabetes Complications ; Diabetes Mellitus ; Diet ; Fasting ; Fruit ; Glucose ; Hemoglobin A, Glycosylated ; Homeostasis ; Hyperglycemia ; In Vitro Techniques ; Insulin ; Insulin Resistance ; Maltose ; Mice ; Olea