The global rise in obesity has become a public health crisis. During the onset of obesity, disrupted catecholamine signals have been described to contribute to excess fat accumulation, however, the molecular and metabolic change of subcutaneous adipose tissue (SAT) upon chronic high-fat feeding has rarely been investigated. Here, we show that chronic high-fat feeding caused a significant decrease in the expression of thermogenic genes and acquisition of partial deleterious features of visceral fat in SAT. Upregulated miR-149-3p was involved in this obesity-induced "visceralization" of SAT via inhibiting PRDM16, a master regulator that promoted SAT thermogenesis. Reduction of miR-149-3p significantly increased PRDM16 expression in SAT, with improved whole-body insulin sensitivity, decreased SAT inflammation, and liver steatosis in high-fat fed mice. These findings provided direct evidence of the anti-obese and anti-diabetic effect of PRDM16 in the obese background for the first time and identified that miR-149-3p could serve as a therapeutic target to protect against diet-induced obesity and metabolic dysfunctions.

It is generally regarded that E-cadherin is downregulated during tumorigenesis via Snail/Slug-mediated E-cadherin transcriptional reduction. However, this transcriptional suppressive mechanism cannot explain the failure of producing E-cadherin protein in metastatic breast cancer cells after overexpressing E-cadherin mRNA. Here we reveal a novel mechanism that E-cadherin is post-transcriptionally regulated by Slug-promoted miR-221, which serves as an additional blocker for E-cadherin expression in metastatic tumor cells. Profiling the predicted E-cadherin-targeting miRNAs in breast cancer tissues and cells showed that miR-221 was abundantly expressed in breast tumor and metastatic MDA-MB-231 cells and its level was significantly higher in breast tumor or MDA-MB-231 cells than in distal non-tumor tissue and low-metastatic MCF-7 cells, respectively. MiR-221, which level inversely correlated with E-cadherin level in breast cancer cells, targeted E-cadherin mRNA open reading frame (ORF) and suppressed E-cadherin protein expression. Depleting or increasing miR-221 level in breast cancer cells induced or decreased E-cadherin protein level, leading to suppressing or promoting tumor cell progression, respectively. Moreover, miR-221 was specifically upregulated by Slug but not Snail. TGF-β treatment enhanced Slug activity and thus increased miR-221 level in MCF-7 cells. In summary, our results provide the first evidence that Slug-upregulated miR-221 promotes breast cancer progression via reducing E-cadherin expression.

Chronic infection with hepatitis B virus (HBV) is a major public health problem. Recently, RNA interfering-based strategy has shown great potential to eradicate HBV infection. In current study, we report the experimental observation of plant-derived artificial microRNAs (amiRNAs) acting as therapeutics in HBsAg-/+ transgenic mice. Two pieces of small silencing RNA sequences, siR471 and siR519, against HBV surface antigen gene (HBsAg) were designed and expressed in lettuce using plant endogenous microRNA biogenesis machinery. Administration of amiRNAs-containing lettuce decoction specifically inhibited the HBsAg gene expression. In long term treatments, the liver injury in HBsAg-/+ transgenic mice were alleviated and no toxicological effects were observed. Compared with synthetic siRNA, feeding amiRNAs at a lower level achieved a similar inhibitory effect on HBsAg expression in mice. These results strongly suggest that employing plant endogenous miRNA biogenesis machinery to generate medicinal siRNAs is a novel way to solve the problems of siRNA stability and reduce the potential side effects of RNAi therapy.

In this study, we investigated the role of serum exosomal miR-222 in obesity-related insulin resistance. Bioinformatics analyses showed that miR-222 levels were significantly upregulated in the white adipose tissue of obese patients with insulin resistance (GSE25402 dataset) and in serum samples from type 2 diabetes mellitus (T2DM) patients (GSE90028 dataset). Moreover, analysis of miRNA expression in adipose tissue-specific Dicer knockout mice (GitHub dataset) and diabetic model mice (GSE81976 and GSE85101 datasets), gonadal white adipose tissue (gWAT) was the main source of serum exosomal miR-222. MiR-222 levels were significantly elevated in the serum, serum exosomes and gWAT of mice fed a high-fat diet (HFD), and there was a corresponding downregulation of IRS1 and phospho-AKT levels in their liver and skeletal muscle tissues, which correlated with impaired insulin sensitivity and glucose intolerance. These effects were abrogated by surgically removing the gWAT from the HFD-fed mice. Thus, gWAT-derived serum exosomal miR-222 appears to promote insulin resistance in the liver and skeletal muscle of HFD-fed obese mice by suppressing IRS1 expression.

Extracellular vesicles (EVs) play an important role in the communication between tissues and cells. However, it is difficult to screen and trace EVs secreted by specific tissues in vivo, which affects the functional study of EVs in certain tissues under pathophysiological conditions. In this study, a Cre-dependent CD63flag-EGFP co-expressed with mCherry protein system expressing mice was constructed, which can be used for the secretion, movement, and sorting of EVs from specific tissues in vivo. This mouse model is an ideal research tool for studying the secretion amount, target tissue, and functional molecule screening of EVs in specific tissues under different pathophysiological conditions. Moreover, it provides a new research method to clarify the mechanism of secreted EVs in the pathogenesis of the disease.

Visceral adiposity is strongly associated with metabolic disease risk, whereas subcutaneous adiposity is comparatively benign. However, their relative physiological importance in energy homeostasis remains unclear. Here, we show that after 24-h fasting, the subcutaneous adipose tissue of mice acquires key properties of visceral fat. During this fast-induced 'visceralization', upregulation of miR-149-3p directly targets PR domain containing 16 (PRDM16), a key coregulatory protein required for the 'browning' of white fat. In cultured inguinal preadipocytes, overexpression of miR-149-3p promotes a visceral-like switch during cell differentiation. Mice deficient in miR-149-3p display an increase in whole-body energy expenditure, with enhanced thermogenesis of inguinal fat. However, a visceral-like adipose phenotype is observed in inguinal depots overexpressing miR-149-3p. These results indicate that in addition to the capacity of 'browning' to defend against hypothermia during cold exposure, the subcutaneous adipose depot is also capable of 'whitening' to preserve energy during fasting, presumably to maintain energy balance, via miR-149-3p-mediated regulation of PRDM16.

RNAi therapy has undergone two stages of development, direct injection of synthetic siRNAs and delivery with artificial vehicles or conjugated ligands; both have not solved the problem of efficient in vivo siRNA delivery. Here, we present a proof-of-principle strategy that reprogrammes host liver with genetic circuits to direct the synthesis and self-assembly of siRNAs into secretory exosomes and facilitate the in vivo delivery of siRNAs through circulating exosomes. By combination of different genetic circuit modules, in vivo assembled siRNAs are systematically distributed to multiple tissues or targeted to specific tissues (e.g., brain), inducing potent target gene silencing in these tissues. The therapeutic value of our strategy is demonstrated by programmed silencing of critical targets associated with various diseases, including EGFR/KRAS in lung cancer, EGFR/TNC in glioblastoma and PTP1B in obesity. Overall, our strategy represents a next generation RNAi therapeutics, which makes RNAi therapy feasible.

Colorectal cancer (CRC) is a major worldwide health problem due to its high prevalence and mortality rate. T-cell intracellular antigen 1 (TIA1) is an important tumor suppressor involved in many aspects of carcinogenesis and cancer development. How TIA1 expression is regulated during CRC development remains to be carefully elucidated.

Exosomes have been widely used in research on the early clinical diagnosis, prognosis and treatment of various cancers due to their features of small size (30-120 nm), non-immunogenicity and ability to cross biological barriers. However, few studies have investigated exosomes involved in metabolic diseases. Early studies have found that adipose tissue can be a source of exosomes regulating metabolism, but the related functions of exosomes secreted by other tissues in the regulation of metabolic diseases have not been determined. In addition, islets were found to be able to secrete miRNA via exosomes, suggesting that islet exosomes may be among the sources of exosomes involved in the regulation of metabolic diseases and that the relevant protein profiles have not been characterized to date. Therefore, identifying the protein contents of pancreatic β cell-derived exosomes would benefit further research investigating the protein functions and mechanisms associated with diabetes-related metabolic diseases. SIGNIFICANCE: Exosomes are emerging tools for investigating metabolic diseases in recent years, but little research has been done. In our work, functional identification of MIN6 cell-derived exosomal proteins and comparative analysis of islet β cell exosomal protein data from different cell lines or from different species revealed that exosomes secreted by islet β cells may be involved in the regulation of glucose metabolism. These results may suggest that intercellular communication induced by exosome transfer among tissues may account for the major reason of diabetic metabolic disorder. In addition, these results may provide a theoretical basis for the study of the physiological and pathological functions of islet β cell exosomes for the future studies.

Secreted microRNAs (miRNAs) are novel endocrine factors that play essential pathological and physiological roles. Here, we report that pancreatic β cell-released exosomal miR-29 family members (miR-29s) regulate hepatic insulin sensitivity and control glucose homeostasis. Cultured pancreatic islets were shown to secrete miR-29s in response to high levels of free fatty acids (FFAs) in vitro. In vivo, high levels of FFAs, promoted by either high-fat diet (HFD) feeding (physiopathological) or fasting (physiological), increased the secretion of miR-29s into plasma. Intravenous administration of exosomal miR-29s attenuated insulin sensitivity. The overexpression of miR-29s in the β cells of transgenic (TG) mice promoted the secretion of miR-29s and inhibited the insulin-mediated suppression of glucose output in the liver. We used selective overexpression of traceable heterogenous mutant miR-29s in β cells to confirm that islet-derived exosomal miR-29s target insulin signalling in the liver and blunt hepatic insulin sensitivity. Moreover, in vivo disruption of miR-29s expression in β cells reversed HFD-induced insulin resistance. In vitro experiments demonstrated that isolated exosomes enriched in miR-29s inhibited insulin signalling in the liver and increased hepatic glucose production. These results unveil a novel β cell-derived secretory signal-exosomal miR-29s-and provide insight into the roles of miR-29s in manipulating glucose homeostasis.

KRAS plays a significant role in the etiology and progression of colorectal cancer (CRC), but the mechanism underlying this process has not been fully elucidated. In this study, we found that the KRAS protein levels were higher in CRC tissues than in the normal adjacent tissues, whereas its mRNA levels varied irregularly, suggesting that a post-transcriptional mechanism is involved in the regulation of KRAS. Then, we performed bioinformatic analyses to search for miRNAs that potentially target KRAS. We predicted and experimentally validated that miR-16 directly recognizes the 3'-UTR of the KRAS transcript and regulates KRAS expression. Furthermore, the in vitro results showed that the repression of KRAS by miR-16 suppressed the proliferation and invasion and induced the apoptosis of CRC cells, and the in vivo results revealed that miR-16 exerted a tumor-suppressive effect by negatively regulating KRAS in xenograft mice. Taken together, our findings provide evidence supporting the role of miR-16 as a tumor suppressor in CRC by targeting KRAS.

miRNAs in circulating extracellular vesicles (EVs) are promising biomarkers for cancer. However, their diagnostic ability for early-stage non-small-cell lung cancer (NSCLC) is not well known. In this study, the circulating EV miRNAs profiling was initially performed in 36 untreated NSCLC patients and 36 healthy controls by TaqMan Low Density Array (TLDA). Subsequently, we performed quantitative reverse-transcription PCR assay (RT-qPCR) validation in several independent cohorts that included 159 NSCLC patients, 120 age/sex-matched healthy controls and 31 benign nodule patients enrolled from three different clinical centres. In addition, 38 cases of NSCLC were analysed before and after surgery. We demonstrated that miR-520c-3p and miR-1274b were significantly and steadily increased in NSCLC patients in comparison with healthy controls and benign nodule patients (P < 0.001) and decreased markedly after tumour resection (P < 0.001). The areas under the curve (AUCs) of the ROC curve of the two-miRNA panel were 0.857 (95% CI, 0813-0.901; P < 0.0001) and 0.845 (95% CI, 0.793-0.896; P < 0.0001) for NSCLC and NSCLC stage I, respectively. Furthermore, the panel was able to differentiate NSCLC from benign nodules with an AUC of 0.823 (95% CI, 0.730-0.915; P < 0.0001). Furthermore, logistic regression analysis revealed the two-miRNA panel as an independent risk factor for NSCLC (OR = 16.128, P < 0.0001). In conclusion, miR-520c-3p and miR-1274b have biomarker potential for early diagnosis of NSCLC in multiple centres.