Cancer cells are well-known for adapting their metabolism to maintain high proliferation rates and survive in unfavorable environments with low oxygen and nutritional deficiency. Metabolic reprogramming most commonly arises from the tumor microenvironment (TME). The events of metabolic pathways include the Warburg effect, shift in Krebs cycle metabolites, and increase rate of oxidative phosphorylation that provides the energy for the development and invasion of cancer cells. The TME and shift in tumor metabolism shows a close relationship through bidirectional signaling pathways between the stromal and tumor cells. Cancer-associated fibroblasts (CAFs) are the main type of stromal cells in the TME and consist of a heterogeneous and plastic population that play key roles in tumor growth and metastatic capacity. Emerging evidence suggests that CAFs act as major regulators in shaping tumor metabolism especially through the dysregulation of several metabolic pathways, including glucose, amino acid, and lipid metabolism. The arrangement of these metabolic switches is believed to shape distinct CAF behavior and change tumor cell behavior by the CAFs. The crosstalk between cancer cells and CAFs is associated with cell metabolic reprogramming that contributes to cancer cell growth, progression, and evasion from cancer therapies. But the mechanism and process of this interaction remain unclear. This review aimed to highlight the metabolic couplings between tumor cells and CAFs. We reviewed the recent literature supporting an important role of CAFs in the regulation of cancer cell metabolism, and the relevant pathways, which may serve as targets for therapeutic interventions. The most prominent biological characteristic of tumor cells is uncontrolled and rapid proliferation. The division of one cell into two requires the doubling of substances such as nucleic acids, amino acids, lipids, and carbohydrates and is the most important metabolic requirement of rapidly proliferating tumor cells, which distinguishes these cells from the vast majority of normal non-cancerous cells. The rapidly proliferating cells must alter their metabolic behavior to achieve the conversion of the highest possible amount of available nutrients into biomass energy, which requires a delicate balance among ATP production, provision of the reducing equivalents, and the synthesis of various biomolecules. In response to factors such as the harsh tumor microenvironment (TME), tumor cells undergo adaptive changes in their metabolic properties, a process referred to as metabolic reprogramming TME, consisting of blood and lymphatic tumor vessels, extracellular matrix (ECM), and non-cancer stromal cells, such as cancer-associated fibroblasts (CAFs), modulates cancer cell growth, progression, and evasion from cancer therapies Associations between CAFs and cancer cells support tumorigenesis in various ways. Besides being a major producer of ECM components, such as collagen and laminins, and remodeling enzymes in vitro and in vivo modeling approaches, have been presented and assessed. Finally, we discussed how reprogrammed CAFs might affect tumor cell metabolic reprogramming and focused on the potential therapeutic strategies targeting molecules involved in the metabolic reprogramming of CAFs.To date, several mechanistic studies have shown the importance of this bioenergetic coupling of tumor and stroma. Therefore, in this review, we focused on the current understanding of the mechanism of cancer cell/CAF metabolic interactions and discussed in detail the activation of different metabolic pathways resulting from these interactions. Detailed metabolic analyses on the tumor cell/CAF crosstalk, including Under physiological conditions, fibroblasts regulate the turnover of ECM, control tissue homeostasis, and participate in wound healing and senescence CAFs act as active participants in the complex metabolism of tumors rather than just the metabolic co-observers of cancer cells Figure . The arrTumor cells perform glycolysis rapidly, even under aerobic conditions, thus increasing glucose uptake and lactate secretion, a phenomenon called the “Warburg effect” or “aerobic glycolysis” In vitro and in vivo (orthotopic xenograft) results showed that hypoxic CAF-derived lactate, which provides metabolic coupling between CAFs and breast cancer cells, promoted breast cancer cell invasion by activating the TGF-β1/p38 MAPK/MMP2/9 signaling axis and fueling the mitochondrial activity in cancer cells It was observed that CAFs proliferated at an unexpectedly slower rate compared to matched NFs, which was in contrast to the proliferation rate of cancer cells, suggesting that increased glycolysis in CAFs did not efficiently increase the proliferation and growth of the CAFs. Bonuccelli et al. found that Cav-1-deficient stromal fibroblasts were sufficient to promote human tumor xenograft growth and angiogenesis. Proteomic analysis of Cav-1-deficient CAFs indicated the upregulation of glycolytic enzymes, such as PKM2 and LDH-B. The researchers speculated that Cav-1-deficient CAFs might contribute to tumor growth and angiogenesis by providing energy-rich metabolites  in a paracrine fashion hi CAFs might promote OSCC cell proliferation hi CAFs is taken up by tumor cells, where it is metabolized to produce NADH, which is then oxidized within the mitochondrial oxidative phosphorylation system (OXPHOS) to generate ATP. Fischi et al. observed that human prostate cancer (PCa) cells induced a high expression of GLUT1 and MCT4 in CAFs, which increased the glucose uptake and lactate output by the CAFs. In contrast, PCa cells, upon contact with CAFs, were reprogrammed for aerobic metabolism, which then exhibited decreased GLUT1 expression and increased lactate uptake via the lactate transporter MCT1 The pancreatic cancer cells expressed significantly higher levels of MCT1, fumarate hydratase (FH), and succinate dehydrogenase (SDH) Amino acids are essential for tumor growth and development, and several studies have demonstrated that CAFs synthesize the amino acids required for tumor cell growth through the TCA cycle k° CAFs can resist glutamine deficiency by directly controlling the stability of activating transcription factor 4 (ATF4) through its p62-mediated polyubiquitination, and thus, maintain PCa cell proliferation k° CAFs. ASNS moderates the synthesis of asparagine from aspartate, while PC mediates the production of oxaloacetate from pyruvate, the route through, which pyruvate enters the TCA cycle. The increased expression of ASNS and PC suggests that the TCA cycle is enhanced, with an increase in the levels of aspartic acid and asparagine. The p62-deficient CAFs produce aspartate and asparagine, which serve as nitrogen sources for the growth of PCa cells, thereby maintaining the survival of PCa cells in a glutamine-deficient environment Besides synthesizing glutamine directly to provide nutrition to tumor cells, CAFs are also capable of producing other amino acids as nitrogen sources. Linares et al. observed that p62 Research suggests that crosstalk exists between human skin squamous carcinoma (HSSC) cells and CAFs in terms of amino acid metabolism These findings demonstrated that CAFs form amino acid metabolic couplings with tumor cells, which enable them to cope with nutrient deprivation in the TME, subsequently allowing the tumors to utilize amino acids efficiently and rapidly to promote tumor growth and development. However, there are many amino acids involved in the metabolic crosstalk between CAFs and tumor cells that have not been determined yet. Further studies are required to decipher the metabolic functions of other amino acids in tumor-CAF interactions.Fatty acids (FAs) are the main building blocks of lipids and can be funneled into various metabolic pathways to synthesize more complex lipid species. FAs contribute to the vast structural diversity of the cellular lipid pool, which in turn serves to regulate several biochemical processes in normal cells In addition to undergoing metabolic reprogramming themselves, CAFs also play an important role in the metabolic reprogramming of tumor cells. CAFs regulate the metabolic remodeling of tumor cells by (I) directly/indirectly exporting nutrients, (II) providing mitochondria, (III) regulating the activity and oxidative properties of metabolic enzymes, and (IV) participating in ECM formation Figure , Table 2Several studies have confirmed that CAFs generate nutrients through catabolic processes and that these nutrients are then absorbed and utilized by tumor cells to maintain tumor growth. Sousa et al. reported that stroma-associated PSCs secrete alanine through activation of autophagy Exosomes are membrane-enclosed vesicles derived from the endosomal system during the formation of multivesicular bodies, with a diameter of ∼30-100 nm 13C isotope-labeling experiments had shown that exosomes could provide amino acids in nutrition-deficient cancer cells through a mechanism similar to macropinocytosis. The researchers performed Gas Chromatography-Mass Spectrometer (GC-MS) and ultrahigh-performance liquid chromatography (UPLC) experiments and confirmed that exosomes in both prostate and pancreatic CAFs contain complete metabolites, including amino acids, lipids, and TCA cycle intermediates, which are extensively used by cancer cells for carbon metabolism during nutritional deficiency or nutritional stress; thus, exosomes promote tumor growth Zhao et al. found that prostate CAF-derived exosomes could inhibit mitochondrial oxidative phosphorylation, thereby increasing glutamine-dependent reductive carboxylation and glycolysis in cancer cells. CAF-CM was collected to isolate exosomes. First, the samples were centrifuged at 2,000 g for 30 min to remove cells and debris; then, they were centrifuged at 10,000g for 60 min at 4 °C. The in vitro studies have shown that PKM can be targeted by miR-330-5p and is controlled by SNHG3 in breast cancer cells. Mechanistically, SNHG3 knockdown in CAF-secreted exosomes suppresses glycolysis metabolism and cell proliferation by the increase in miR-330-5p and decrease in PKM expression in tumor cells. SNHG3 functions as a miR-330-5p sponge to upregulate PKM expression, inhibit mitochondrial oxidative phosphorylation, increase glycolysis carboxylation, and enhance breast tumor cell proliferation Besides directly delivering nutrients, exosomes derived from CAFs can upregulate glycolytic and glutamate-dependent reductive carboxylation capacity in cancer cells through the transfer of noncoding RNAs and substrates. Li et al. demonstrated that breast cancer cells reprogram the metabolic pathways after taking the exosomes secreted from CAFs in vivo and in vitroAltered mitochondrial function is reported to significantly regulate tumor metabolism Interestingly, the CAF-derived exosomes also provide the mitochondrial genome to the tumor cells. In drug-resistant breast cancer, the intact mitochondrial genome  of CAFs was observed to be packaged in the exosomes and transferred to breast cancer cells. The CAF-derived mtDNA can be expressed in an intact form within the breast cancer cells and can increase the levels of tumor oxidative phosphorylation and mitochondrial metabolism, which further enables tumor drug resistance and enhances the self-renewal potential of tumor stem cells in vitro and in vivo were associated with an elevated expression of the chemokine CXCL14. CXCL14-high CAFs mediated upregulation of LINC00092 in ovarian cancer cells, the levels of which also correlated with poor prognosis in patients. Mechanistic studies have shown that LINC00092 binds to a glycolytic enzyme, the fructose-2,6-biphosphatase PFKFB2, thereby promoting metastasis by altering glycolysis in cancer cells and sustaining the local supportive function of CAFs In 2015, our research team had observed that thymic fibroblast specific protein-1 (FSP-1)-positive fibroblasts released high amounts of IL-6, fibroblast growth factor 7 (FGF7), and S100A4 in the culture medium, which are crucial for the maintenance and regeneration of thymic epithelial cells Besides affecting the key enzymes of metabolic pathways, CAFs can also influence the metabolism-related signaling pathways in tumor cells. Tommelein et al. reported that preoperative radiotherapy (RT) for CRC elicits secretion of insulin-like growth factor 1 (IGF-1) from CAFs 2O2 production by CAFs was associated with impaired TGFβ signaling that led to the suppression of the antioxidant enzyme glutathione peroxidase 1 (GPX1). This consequently increased ROS levels and fueled tumor growth The redox state of tumors is often imbalanced, as shown by the increased levels of reactive oxygen species (ROS), which are important for both drug resistance and immune tolerance. Broekgaarden et al. reported that in an orthotopic xenograft model of PDAC, the tumors co-implanted with PDAC cells and CAFs exhibited an increased oxidative state and significantly increased resistance to oxaliplatin, compared to the tumors injected only with PDAC cells; the half-maximal inhibitory concentration (IC50) increased by 2.4-fold Tumor tissues contain a large number of ECM molecules, which besides maintaining the mechanical tension of the matrix, also communicate with cells via cell membrane surface receptor integrins Besides causing metabolic dysregulation of local tissues, cancer is also associated with altered metabolism in the host Increased systemic levels of pro-inflammatory cytokines are considered to be strongly linked to host metabolic disturbances associated with cancer progression Besides focusing on the tumor-supporting role of CAFs, we also need to pay attention to some “alternative” studies. Three published papers simultaneously mentioned the protective effect of CAFs against PDAC, which may be related to the inhibition of pro-cachectic mechanisms Tumor cells must be capable of thriving in a nutrient-deprived tumor microenvironment to survive, a process in which CAFs assist the tumors to cope with the nutrient deprivation and maintain tumor growth, metastasis, and drug resistance in vivo studies have demonstrated that inhibiting the exchange of substances between CAFs and tumor cells, such as by the lactate shuttle An increasing number of studies are focusing on tumor metabolism, and within this area of research, the regulation of tumor metabolic reprogramming by CAFs is an important direction with a strong academic research value in vivo and in vitro conditions. Therefore, it is imperative to understand the impact of tumor heterogeneity and the failure to recognize the nonlinear relationship between micronutrients and cancer in fundamental research and clinical trials.However, most of these studies are currently at a fundamental level, and there is a lack of reported clinical studies. Moreover, several obstacles need to be overcome, such as the limited number of methods available to measure the metabolism of CAFs, the uncertainty in metabolic heterogeneity, and the lack of metabolic similarity under


Hepatosteatosis is the earliest stage in the pathogenesis of nonalcoholic fatty (NAFLD) and alcoholic liver disease (ALD). As NAFLD is affecting 10–24% of the general population and approximately 70% of obese patients, it carries a large economic burden and is becoming a major reason for liver transplantation worldwide. ALD is a major cause of morbidity and mortality causing 50% of liver cirrhosis and 10% of liver cancer related death. Increasing evidence has accumulated that gut-derived factors play a crucial role in the development and progression of chronic liver diseases.A selective literature search was conducted in Medline and PubMed, using the terms “nonalcoholic fatty liver disease,” “alcoholic liver disease,” “lipopolysaccharide,” “gut barrier,” and “microbiome.”Gut dysbiosis and gut barrier dysfunction both contribute to chronic liver disease by abnormal regulation of the gut-liver axis. Thereby, gut-derived lipopolysaccharides (LPS) are a key factor in inducing the inflammatory response of liver tissue. The review further underlines that endotoxemia is observed in both NAFLD and ALD patients. LPS plays an important role in conducting liver damage through the LPS-TLR4 signaling pathway. Treatments targeting the gut microbiome and the gut barrier such as fecal microbiota transplantation (FMT), probiotics, prebiotics, synbiotics, and intestinal alkaline phosphatase (IAP) represent potential treatment modalities for NAFLD and ALD.The gut-liver axis plays an important role in the development of liver disease. Treatments targeting the gut microbiome and the gut barrier have shown beneficial effects in attenuating liver inflammation and need to be further investigated. Alcohol intake can lead to changes in gut microbiota composition, even before the onset of liver disease.9Alcoholic liver disease (ALD) is a major cause of morbidity and mortality among people who abuse alcohol.10 In humans, the gut microbiota is the major source of LPS.11 LPS is generally known for its role in the induction of sepsis, septic shock, and multiple organ failure.12 Recently, LPS is found to be related to several other diseases, especially to metabolic disorders such as type 2 diabetes mellitus and atherosclerosis.14 Furthermore, the role of LPS in neurologic and mental disorders like Alzheimer’s disease and autism has been well described,16 linking gut microbiota to the homeostasis of the entire body.Lipopolysaccharide (LPS), also known as endotoxin, is a component of the outer cell wall of Gram-negative bacteria.17 Bacterial overgrowth in the blind loop has been considered to be responsible for liver damage. Today, the human intestinal microbiota has emerged as an important mediator of the development and progression of chronic liver diseases.The connection between gut microbiota and chronic liver disease has been first noticed in the 1980s when NASH was encountered as a common complication of jejunoileal bypass surgery for morbid obesity that could be reversed by treatment with metronidazole.This manuscript briefly reviews the role of gut-derived LPS in the development and progression of NAFLD and ALD. Gut barrier dysfunction and dysbiosis are highlighted as they are the two major mechanisms of endotoxemia as well as potential therapeutic targets.18The intestinal barrier serves as a physical and functional barrier deterring translocation of potentially harmful luminal antigens into circulation. The four layers of the intestinal barrier include luminal intestinal alkaline phosphatase (IAP) released from the intestinal epithelial cells, surface mucus, epithelial layer, and immune defense. The epithelial cell layer and the mucin layer constitute the physical barrier; IAP and antibacterial proteins secreted by Paneth cells represent the functional barrier.19 The tight junctions (TJs) constitute the major determinant of the intestinal physical barrier and can prevent the paracellular passage of large molecules through the epithelium.20IAP dephosphorylates substrates such as LPS and other pathogen-associated molecular patterns. Its role in reducing local intestinal inflammation and maintaining gut barrier function is reviewed later. The intestinal mucosal layer represents the first physical barrier. The small intestine harbors a single, tightly attached mucus layer, whereas in the colon, the mucus is composed of two layers: the essentially sterile inner layer which is firmly attached to the epithelial cells and the outer layer which contains commensal bacteria that prevent the entry of pathogenic bacteria. The epithelium is constituted of a single layer of different intestinal epithelial cells (IECs), and cells within the epithelial layer are sealed by tight junction proteins including claudins, zonula occludens-1 (ZO-1), and occluding, preventing paracellular transport.21 Many factors can alter intestinal permeability, such as diet, alcohol intake, medication, and physiological factors such as age and stress.24 Changes in intestinal microbiota composition can influence intestinal permeability, as a study showing that a high-fat diet changed the gut microbiota content and could increase intestinal permeability. More importantly, this effect was completely restored by antibiotic treatment.25 And vice versa, probiotic bacteria and probiotic mixture are shown to have beneficial effects in reestablishing intestinal homeostasis and preserving epithelial barrier function.26 Bacterial metabolites, such as short-chain fatty acids (SCFAs), are also reported to play an important role in maintaining both intestinal immune functions and regulating gut barrier functions.27Disruption of the intestinal barrier can cause LPS translocation and cause endotoxemia in systemic circulation and chronic liver inflammation. The permeability of the epithelium is determined by the composition and abundance of different components of the TJs.28 Translocation of LPS can occur in physiological states.29 Under pathological conditions when there is an increase in gut barrier permeability, paracellular pathway of LPS is increased. The liver is the first organ in the body to encounter gut-derived bacteria and pathogen associated molecular patterns (PAMPs). Chronic exposure to increased levels of PAMPs has been linked to liver diseases.30LPS translocate the intestinal barrier mainly through the transcellular pathway, and chylomicrons can also bind and facilitate the absorption of LPS.31 The portal venous system sits at the interface between the host and the inflammatory mediators that exist within the gut. The most important gut-derived inflammatory mediator, LPS, enters the liver through the portal vein and is detoxified in the liver. Only few LPS passes through the gut barrier and finally arrives in the liver under physiologic conditions. However, small intestinal bacteria overgrowth (SIBO) or an increase in intestinal permeability leads to translocation of bacteria and its by-products such as LPS. After arriving in the liver, LPS is taken up by hepatocytes and Kupffer cells and is excreted into the bile duct in further process.32 In patients with chronic liver disease, high LPS portal/peripheral concentrations have been observed.33 LPS binds to LPS-binding protein (LBP), and the LBP-LPS complex is transferred to membrane bound or soluble cluster of differentiation 14 (CD14), thereby specifically binds to toll-like receptor 4 (TLR4) and induces the interaction of TLR4 with adaptor molecule myeloid differentiation factor 88 (MyD88). MyD88 further activates downstream mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-kB). Stimulation of the LPS-TLR4 signaling pathway eventually leads to the release of proinflammatory mediators like tumor necrosis factor-α (TNF-α) and interleukin 6 (IL-6).34 LPS also mediates signaling through MyD88-independent pathway, but the activation of MAPK and NF-kB occurs in a delayed manner.35 In both ALD and NAFLD progression, TLR4 signaling is considered a key pathway, and, very interestingly, it is reported that mice deficient in TLR4 are resistant to both alcohol-induced liver injury and NAFLD.37As the gut is considered to be the first barrier against bacteria, the liver is the second barrier based on the fact that the liver and the gut share both anatomical and functional relations and proximities; their close interaction is also described as the gut-liver axis.32 In normal liver, hepatic cells express minimal TLRs; therefore, the liver has a high tolerance to TLR ligands. Hepatocytes directly clear LPS as data showed that fluorescence was revealed in hepatocytes 5 min after injection of fluorescein isothiocyanate (FITC)-LPS into the portal vein and then rapidly secreted into the bile.38 LPS promotes TNF-α production in Kupffer cells. The inflammatory mediator TNF-α is considered a central mediator in the pathogenesis of both ALD and NAFLD.39 Despite the fact that Kupffer cells are the main targets of LPS in the liver, it is the HSCs to promote TLR4-dependent fibrosis.40 LPS can activate HSCs in vitro and in vivo, and Kupffer cells strongly enhance this process by producing transforming growth factor beta (TGF-β) and increasing the sensitivity of HSCs to TGF-β. Recent studies have pointed out that platelets can also play a role in liver injury as biopsy results showed that the number of platelets is associated with disease severity in NAFLD.33 Platelets passing through hepatic sinusoids can be activated by LPS and the number of TLR4+ platelets is positively correlated with serum LPS level in NAFLD patients, suggesting that LPS may activate platelets through TLR4 signaling pathway. LPS is capable of activating platelets via TLR4-mediated over-production of eicosanoids, and mice given aspirin showed a decrease of liver fibrosis.42TLR4 is expressed on all types of liver cells, including Kupffer cells, hepatocytes, hepatic stellate cells (HSCs), and also cholangiocytes. Its expression on these cells is correlated with activation of fibrogenic cells and the stage of fibrosis.43 indicating that there may be a common pathway for liver injury in both diseases. The hallmark of NAFLD and ALD is the excessive fat accumulation in the hepatocytes, which may be an isolated event (simple steatosis) or accompanied by evidence of inflammation and cell injury with or without fibrosis (NASH and ASH)44. The histological changes in ALD or NAFLD are lobulocentric  but not portal-based,45 including hepatocyte ballooning, Mallory bodies, zone 3 inflammation, and perisinusoidal fibrosis. Recent studies have identified the importance of portal fibrosis in predicting the subgroup of NAFLD patients that develop progressive liver disease and liver-related mortality.46 Portal inflammation is associated with portal-based changes, such as ductular reaction (DR), a reactive lesion at the portal tract interface comprising small biliary ductules with an accompanying complex of stroma and inflammatory cells.47 The underlying mechanism of DR is the activation of a secondary proliferative pathway of hepatic progenitor cells (HPCs) during the replacement of necrotic and apoptotic hepatocytes.48 Under regular conditions, the primary pathway of liver regeneration is maintained by the replication of adjacent hepatocytes within the lobules. With an insult of toxins, viral infection, alcohol, etc., the primary pathway is blocked and replaced by the secondary pathway: HPCs proliferate and differentiate into hepatocytes and bile ductal epithelia; the by-product is the DR . The study found that the keratin 23 (KRT23) gene, which is expressed in the ductular reaction cells, is the most upregulated gene in AH compared to NASH and normal livers. They discovered that LPS administration markedly induced KRT23 expression in mice. This effect can be attenuated in TLR4-deficient mice, further indicating that the LPS-TLR4 pathway mediates the development of ductular reaction in chronic liver injury.Odena et al.32 provided evidence that serum LPS levels correlated with portal/interface inflammation, the activity of portal/septal myofibroblasts, and fibrosis in NAFLD patients. Using immunohistochemistry, TLR4 expression was observed in hepatic progenitor cells, biliary cells, and portal/septal macrophages. TLR4-positive hepatic progenitor cells and bile ducts/ductules correlate with portal/interface inflammation, activation of fibrogenic cells, and fibrosis, implicating that the LPS-TLR4 pathway is associated with inflammation and fibrosis progression in NAFLD.Another study by Vespasiani-Gentilucci et al.50 The etiology of NAFLD is previously referred to as a “two-hit” hypothesis:51 The first hit involves lipid accumulation in the hepatocytes, making the liver more vulnerable to toxins. Further insults like ethanol and LPS can act as the second hit and amplify the initial stress, causing oxidative stress and inflammatory response within the liver, resulting in the development of steatohepatitis  may occur parallelly, leading to liver inflammation. Among these hits, gut-derived factors appear to play a central role. Although traditional views consider that simple steatosis and NASH are consecutive changing processes, emerging evidence indicates that these two diseases can arise as independent conditions.54 Studies have found that the patients with simple steatosis will not always develop NASH and that the two diseases have a huge difference in prognosis in long term follow-up. This evidence further demonstrates that steatosis may not always precede inflammation.NAFLD is the liver manifestation of metabolic syndrome and is characterized by massive ectopic triglyceride accumulation in the liver in the absence of any other liver disease or significant alcohol consumption.s Figure . The “tw55 IR is thought to contribute to NAFLD by enhancing lipolysis of the adipose tissue, resulting in the increase of free fatty acids flux into the liver.56 High-fat, high cholesterol diets can result in endotoxemia and low-grade inflammation in both human and animal models.58 Lifestyle modifications such as diet, physical exercise, and weight loss are advocated. The Mediterranean diet characterized by reduced carbohydrate intake and increased monounsaturated and omega-3 fatty acid intake has a beneficial effect in overweight patients with NAFLD.59 A previous study showed that even short-term diet modification could reduce liver steatosis and steatohepatitis.60Insulin resistance (IR) has been recognized as a major factor in the development of NAFLD.61 Gut dysbiosis is causative for the enhanced secretion of LPS and the resulting inflammation in NAFLD development.62 Generally, dysbiosis is defined as an imbalance or alteration in the microbiota that can have an unfavorable effect on the host.63 A previous study investigated taxonomic compositions of gut microbiota within the spectrum of NALFD lesions and found that more serious NAFLD lesions (NASH and significant fibrosis) associate with gut dysbiosis.64 The study also found that increased levels of Bacteroides were independently associated with NASH and increased Ruminococcus abundance with fibrosis, linking these two taxa of bacteria to the severity of NAFLD.Endotoxin plasma level is significantly higher in NAFLD patients, and it seems that endotoxin level is associated with the severity of hepatic steatosis.65 It has been shown that microbiota play an important role in nutrition intake and patients with obesity or metabolic disorder appear to have an intestinal microbiota signature that can harvest more energy than their healthy counterparts.66 Changes in abundance and diversity of the gut microbiota in NAFLD have also been characterized. Though many studies have reported conflicting results of the microbiome signatures in NAFLD, an increase in Firmicutes and a reduction in Bacteroidetes is found in most studies.67 Thus, a higher Firmicutes/Bacteroidetes ratio is frequently cited as a hallmark of obesity.68 Furthermore, the amount of ethanol-producing bacteria  was reported to be increased in NAFLD, which may cause increased intestinal permeability and increased translocation of endotoxins from the intestinal lumen to the portal blood.69 The impact of microbiota on liver disease is further supported by experiments during which bacterial transfer from the human gut to germ-free (GF) mice resulted in metabolic diseases71The gut microbiota composition of an individual is a kind of a fingerprint highly influenced by the type of diet.72 found that LPS per se can cause an increase in intestinal epithelial tight junction permeability in vitro and in vivo. This effect was mediated by an increase in enterocyte TLR4 expression and a TLR4-dependent increase in membrane CD14 expression. Studies have shown that physiologically relevant concentrations of LPS (0 to 2000 pg/mL) cause an increase in intestinal epithelial TJ permeability.74 These findings indicate that LPS is an important pathogenic factor in the intestinal inflammatory process.Guo and colleguesThe gut microbiota is influenced by various factors, including the genetic background of the host, type of diet, age, and medication. How these factors exactly affect gut microbiota composition and function is not fully understood with several studies presenting contradictory data. The regulation of gut microbiota provides a new insight into the treatment of NAFLD.75 A previous study showed that intestinal hyperpermeability occurs only in alcoholics with ALD and not in those without liver disease.76 Furthermore, chronic alcohol administration has been shown to increase gut-derived endotoxin levels in the portal circulation.77 This indicates that alcohol plays a crucial role in promoting intestinal hyperpermeability resulting in endotoxemia, systemic inflammation, and liver damage. Alcohol consumption destroys the integrity of the intestinal barrier, disturbs the gut microbiota, and is associated with an increase in the abundance of endotoxin-producing bacterial types.78 Metagenomic analysis of the intestinal microbiome of individuals with a history of chronic alcohol abuse has revealed reduced bacterial diversity and a lower proportion of Bacteroidaceae and probiotic bacteria such as Lactobacillus80. Patients with alcoholic cirrhosis have an increased relative abundance of Enterobacteriaceae.81 Alcohol consumption affects bacterial composition within specific phyla. A study by Llopis et al.82 found that mice harboring the intestinal microbiota from patients with severe alcoholic hepatitis developed more severe liver inflammation, indicating that individual susceptibility to ALD is substantially driven by the intestinal microbiome.ALD is associated with high morbidity and mortality rates. Most chronic heavy drinkers develop steatosis, but only 35% develop advanced liver disease.83 Endotoxemia in ALD was first recognized by the detection of antibodies against Escherichia coli in the plasma of patients with ALD.84 Humans and animals with chronic alcohol consumption develop a “leaky gut,” as evidenced by higher levels of plasma endotoxin. The plasma endotoxin levels are also associated with the severity of disease.85 Alcohol damages specific components of the intestinal barrier such as proteins involved in innate antibacterial defense. Animal experiments have shown that chronic exposure to ethanol can cause gut-barrier disruption featured by a decrease in tight junction protein ZO-1 and occludin expression.87Both chronic alcohol consumption and acute alcohol intake (binge drinking) can impair the intestinal barrier and increase serum levels of bacterial products.88 The underlying mechanism includes the dissociation of ZO-1 from the junctions in acetaldehyde-treated cell monolayer, thus leading to disrupted TJ structure and the increase in paracellular permeability.89 The study further revealed that acetaldehyde increases tyrosine phosphorylation of ZO-1, E-cadherin, and β-catenin, resulting in disruption of the TJs.Besides ethanol, the by-product acetaldehyde from the metabolization of alcohol by gut microbiota appears to play a crucial role in gut barrier dysfunction. At concentrations ranging from 99 to 760μM, acetaldehyde increases the paracellular permeability of Caco-2 cell monolayer.90 A study by Xie et al.91 showed that ethanol consumption could lead to a substantial decrease in taurine-conjugated (hydrophilic and less toxic) bile acids, resulting in impaired lipid emulsification and liver steatosis in mice.Chronic alcohol consumption is also associated with changes in bile acid profiles. ALD patients have an increased secondary bile acid formation.While simple steatosis is considered a “benign disease,” treatment of NAFLD and ALD should focus on NASH and ASH. Currently, there is no targeted and thus effective drug therapy for the treatment of NASH or ASH. Especially, the long-term intake of traditional treatments for chronic liver diseases such as antibiotics and corticosteroids is associated with various side effects for patients. There is accumulating evidence that the interplay between the gut microbiota and the liver is critical in the pathogenesis of chronic liver disease. Hence, there is a large interest in modulating the microbial community to achieve a therapeutic effect or even reverse liver fibrosis. The manipulation of the gut microbiome by fecal microbiota transplantation (FMT), probiotics, prebiotics, and synbiotics was shown to have a beneficial effect in improving liver phenotype in patients with chronic liver disease. IAP as a naturally occurring brush boarder enzyme that detoxifies LPS and preserves microbial homeostasis and gut barrier integrity appears to be a promising candidate for treatment of liver fibrosis in patients with NAFLD or ALD.Clostridium difficile infection (rCDI) for years93 and possesses potential in treating gastrointestinal as well as extraintestinal diseases.This procedure involves the transfer of processed feces to restore a “healthy microbiome.” FMT has been successfully used to treat recurrent 94 The therapeutic effect may be caused by an increase in “beneficial” gut microbiota, improving the tight junction of small intestinal and lowering the LPS levels. Ferrere et al.95 performed FMT from alcohol-resistant donor mice  to alcohol-sensitive receiver mice  and found that FMT protected the alcohol-sensitive mice from alcohol-induced depletion of Bacteroides. FMT treatment was also found to prevent steatosis in alcohol-fed mice, indicating the beneficial effect of FMT in preventing ALD development. In a small-scale pilot study conducted by Philips et al.,96 8 patients with steroid-ineligible severe alcoholic hepatitis received FMT treatment from healthy donors. The results showed that indices of liver disease severity, including ascites, hepatic encephalopathy, and mean bilirubin, significantly improved after FMT. FMT treatment also showed an effect on modulation of gut microbiota and improved prognosis in these patients. In a recent randomized trial, twenty patients with liver cirrhosis were randomly given FMT capsules or placebo. Patients receiving FMT capsules showed an improved gut microbial function and significantly reduced systemic inflammation markers (IL-6 and LBP).97FMT has shown an effect in alleviating high-fat diet-induced steatohepatitis in mice.FMT contributes to restore a balanced gut microbiota composition and has emerged as a therapeutic option for chronic liver disease. Although FMT shows its beneficial effect in animal models and clinical trials of non-alcoholic and alcoholic liver steatosis, it has not been approved for clinical use. Further research is needed to verify the safety and effectiveness of FMT in chronic liver disease.98 They exhibit characteristics including tolerance to gastrointestinal conditions, ability to adhere to the gastrointestinal mucosa and competitive exclusion of pathogens.99 The strains most frequently used as probiotic bacteria belong to the Bifidobacterium and Lactobacillus genera and are widely used in yogurts and other dairy products.100 Probiotics are generally considered to have a beneficial effect on the human intestine. Induction of probiotics may control the growth of pathologic organisms. Studies have proven that probiotics are efficient in treating various diseases such as antibiotic-associated diarrhea (ADD), inflammatory bowel diseases (IBD), and neuropsychiatric disorders. 103Probiotics are defined by the Food and Agriculture Organization (FAO)/World Health Organization (WHO) as living microorganisms that confer a health benefit on the host when administered in adequate amounts.104 Plaza-Diaz et al.105 demonstrated the reduction in liver steatosis of obese mice fed certain probiotic strains. This effect was associated with lower serum LPS levels as matched trends were observed in LPS serum concentration and liver TG content.Probiotics have been proven to lower the plasma triglyceride (TG) levels in animal models of metabolic syndrome.108 Probiotic treatment has been shown to play a role in improving liver function. The intake of probiotics leads to reduced levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST), glutamine transferase (GGT), total cholesterol (TC), TG, and improved liver histological marker such as NAS (NAFLD Activity Score).107 Probiotic supplementation is also able to improve biomarkers of inflammation  and steatosis .109Clinical trials have confirmed the beneficial effects of probiotics in nonalcoholic liver diseases both in adults and children.110, Bifidobacterium bifidum and Lactobacillus plantarum 8PA3 were randomly given to patients diagnosed with alcoholic psychosis. The treatment led to the alterations in bowel flora in alcoholic patients and liver enzyme reduction, indicating that probiotics can improve alcohol-induced liver injury. Other randomized-controlled trials of Lactobacillus casei supplements in patients with alcoholic liver injury support this finding.111Similar to NAFLD, patients with alcoholic liver disease can also benefit from a probiotic treatment. In a Russian pilot study112In general, the mechanisms by which probiotics prevent chronic liver injury include: reducing gut-derived microbial LPS by restoring the bowel flora, repairing the intestinal mucosa and barrier function, modulation of the immune system, and reducing inflammatory cytokine levels.113 Main prebiotics include primarily short- and long-chain fructans (fructo-oligosaccharides (FOS) and inulin), galacto-oligosaccharides (GOS), and lactulose.114 These substances possess features including non-digestibility, fermentation by intestinal microflora, and selective stimulation of growth and activity of intestinal bacteria.115A prebiotic is a non-viable food component that can confer a health benefit on the host that is based on the modulation of the intestinal microbiota.116 Prebiotics are used mostly as a selective medium for the growth of a probiotic strain, and Lactobacilli and Bifidobacteria are the usual target genera for prebiotics. Large bundles of studies have shown that prebiotics are able to increase the composition and/or activity of Lactobacillus and Bifidobacterium populations.120 In lipid metabolism, prebiotics exhibit serum or hepatic lipid-lowering properties, and are now considered a potential dietary adjunct in reducing the risks of cardiovascular diseases (CVD) with minimal side effects121. A randomized placebo-controlled trial carried out in Canada showed that, after 16 weeks of consumption of oligofructose-enriched inulin, there was a significant decrease in body weight z-score, percent body fat, and percent trunk fat and serum level of IL-6 in healthy children with overweight or obesity compared to the placebo control group.122 The study also observed a significantly higher abundance of Bifidobacterium spp. in the prebiotic consumption group, indicating that prebiotic consumption could selectively alter gut microbiota.Prebiotics have beneficial effects on the gastrointestinal tract, which include prevention of pathogen damage or immune system modulation, improvement of gut barrier function, reduction in the pathogenic bacteria population, and the production of SCFAs.124 However, the effect of inulin in human trials remains controversial. Chambers et al.125 explored the effects of dietary supplementation with inulin in adults with NAFLD and found that inulin consumed at 20 g/d increased intrahepatocellular lipid (IHCL). They speculated that the acetate derived from colonic fermentation of inulin could provide an additional lipogenic substrate to the liver.The prebiotic inulin is proven to be effective in preventing NAFLD in animal experiments.115 It is known that probiotics are active in the small and large intestine and the effect of a prebiotic is mainly in the colon. The combination of the two may have a synergistic effect.126 The intake of synbiotics has been demonstrated to modify the composition of the microbiota, thus protecting against inflammation and hepatocyte damage.127 A recent meta-analysis involving 15 randomized clinical trials in which 8 studies about synbiotics were included showed that synbiotics supplementation can improve TC, TG, high-density lipoprotein (HDL), and low-density lipoprotein (LDL) in patients with NAFLD, indicating that synbiotics could improve lipid profiles in those patients.128 Among these studies, Malaguarnera et al.127 conducted a randomized clinical trial in which 66 NASH patients were enrolled and randomly given Bifidobacterium longum with fructo-oligosaccharides or placebo. The results showed that synbiotic treatment improved liver histology in NASH patients and had a beneficial effect in reducing inflammation markers like C-reactive protein (CRP) and TNF-α. These effects may be due to reduced LPS exposure to the liver.A synbiotic is a mixture of 1 or more probiotics and 1 or more prebiotics that beneficially affect the host by promoting the survival and colonization of the live microbes in the gastrointestinal tract.130 Synbiotic supplementation was also shown to decrease serum LPS levels in high-risk alcoholic participants and in patients with alcohol-related cirrhosis.132Though studies of the relationship between synbiotics and ALD are much lesser, the beneficial effect of synbiotics in attenuating chronic alcohol intake induced liver injury is observed in mice.133 IAP is secreted bilaterally and is also released in small amounts into the blood.134 IAP plays an important anti-inflammatory role by dephosphorylating potentially pro-inflammatory ligands such as adenosine triphosphate (ATP), uridine diphosphate (UDP), unmethylated cytosine-guanosine dinucleotides (CpG), and LPS.135 Luminal IAP can prevent and reduce intestinal inflammation and bacterial translocation and is considered the first layer of the intestinal barrier.18 The anti-inflammatory effect makes it a potential treatment for various diseases. Previous studies had found decreased levels of IAP protein expression in inflamed colonic mucosa in children with IBD and decreased IAP mRNA levels in inflamed tissue of adults with IBD.137 Exogenous IAP supplementation showed a beneficial effect in alleviating inflammation in a rat model of IBD.138 In a phase II trial, intravenous injection of IAP improved renal function in patients with severe sepsis and septic shock.139 And another phase II trial demonstrated that administration of bovine IAP could reduce post-surgical inflammatory response in patients undergoing coronary artery bypass grafting (CABG).140IAP is ubiquitously expressed by enterocytes in the proximal small intestine and exists in high concentrations within luminal vesicles secreted by enterocytes on the brush border of the microvilli.3− secretion and localized extracellular pH (pH0).141 Liu et al.142 demonstrated that IAP can directly regulate TJ protein levels as its gene depletion in mouse embryonic fibroblasts resulted in significantly lower levels of ZO-1, ZO-2, and occludin expression, and IAP overexpression in Caco-2 and T84 cells resulted in increases in mRNA levels of ZO-1 and ZO-2  mice and found dramatically fewer microbes in their stools compared with wild-type (WT) mice when fed a high-fat diet. Moreover, oral supplementation of calf IAP (cIAP) promoted the restoration of the normal gut microbiota following antibiotic treatment. Kühn et al.145 observed the beneficial effect of IAP supplementation in preserving the homeostasis of gut microbiota during aging in mice. IAP has been shown to prevent chronic liver disease in 2 different mice models. Liu et al. 146 demonstrated that fecal IAP activity decreases in humans with liver cirrhosis and oral supplement of IAP attenuated liver fibrosis in mice. Hamarneh et al.147 demonstrated that pretreatment with IAP attenuated the development of alcohol-induced fatty liver, decreased hepatic pro-inflammatory cytokines, as well as serum LPS levels, and prevented alcohol-induced gut barrier dysfunction in mice, indicating that oral IAP supplementation could present a novel therapy to prevent alcoholic-related liver disease. Though the beneficial effect of IAP in chronic liver diseases and maintaining gut barrier function has been observed in various preclinical studies, the therapeutic value of IAP in patients with chronic liver disease is not known.IAP also plays a crucial role in regulating the gut microbiota. Malo et al.Accumulating evidence has pointed out the importance of the gut-liver axis in the development of liver disease. Translocation of bacterial by-products such as LPS results in chronic inflammation in the liver. Treatments targeting the gut microbiome such as FMT, probiotics, prebiotics, and synbiotics have shown an effect in attenuating liver inflammation. IAP possessing an anti-inflammatory effect by dephosphorylating LPS and maintaining gut barrier function is also showing therapeutic potential. Current evidence demonstrates that maintaining gut barrier integrity and microbiome homeostasis is of great significance in treating liver diseases.


Various pollutants of different sizes are directly  and indirectly  threatening our water health and safety. To cope with this matter, multifaceted approaches are required for advanced wastewater treatment more efficiently. Wastewater treatment using mixed matrix membranes (MMMs) could provide an excellent alternative since it could play two roles in pollutant removal by covering adsorption and size exclusion of water contaminants simultaneously. This paper provides an overview of the research progresses and trends on the emerging materials used to prepare MMMs for pollutant removal from water in the recent five years. The transition of the research trend was investigated, and the most preferred materials to prepare MMMs were weighed up based on the research trend. Various application examples where each emerging material was used have been introduced along with specific mechanisms underlying how the better performance was realized. Lastly, the perspective section addresses how to further improve the removal efficiency of pollutants in an aqueous phase, where we could find a niche to spot new materials to develop environmentally friendly MMMs, and where we could further apply MMMs. Water contamination by organic and inorganic pollutants emitted from anthropogenic activities is a serious global problem found everywhere regardless of whether the country is developed or developing. In most developing countries, wastewater treatment plants consist only of physical (primary) and biological (secondary) treatments due to the lack of proper infrastructure for tertiary treatment in many cases . For thaMembrane-based technologies used for water treatment can be an excellent alternative to handle wastewater including both conventional and emerging pollutants as they can provide technological  and economic  advantages over other water treatment processes ,9,10. HoAs a promising alternative, nanomaterial-based membranes can be effective, as nanomaterials are highly likely to offer distinct advantages over conventional polymeric materials used for membrane fabrication. Particularly, two-dimensional (2D) nanomaterials such as graphene oxide (GO) and Mxenes have shown great potentials when used in the form of laminate membranes in the filtration processes. In detail, they feature ultra-fast transport of water molecules, precise molecular sieving, and tunable physicochemical properties ,12, and 2SO4/HNO3 for GO [2 [Specifically, 2D nanomaterials typically require a considerable amount of acid or hard-to-handle chemicals . A volu3 for GO ,15 or reor GO [2 ). HoweveIn this regard, mixed matrix membranes (MMMs) comprising polymer matrix-containing fillers can be a viable alternative to overcome the above-mentioned limitations of laminate membranes imposed by the inherent properties of 2D nanomaterials. MMMs can confer additional functions . In addition, MMMs for gas separation were also excluded, given that this paper focuses on pollutant removal in water. Although the literature survey was performed for the research papers published since 2010, a total of 55 papers were found to be published from 2016 to the first quarter of 2021. This result means that the topic related to MMMs for pollutant removal has only recently begun to be studied and is still at an early stage. A more important point is the increasing trend in the number of publications A. Three Another point worthy of note is the most preferred additive type to prepare MMMs for pollutant removal. To be specific, inorganic nanomaterials have been most frequently used as an additive throughout the whole period, which was evidenced by MMMs are heterogeneous membranes prepared by incorporating inorganic fillers with high selectivity into a polymer matrix featuring excellent mechanical properties and processability. More broadly, an MMM can be defined as a membrane consisting of a polymer matrix containing “homogeneously dispersed fillers (not limited to inorganic fillers)” with different selectivity and transport properties as described in the literature . Note thGetting back to the point, as an MMM can be broadly defined as above, a great variety of MMMs can be prepared by combining various kinds of polymers and fillers. Regardless of types of polymers and fillers, MMMs are typically prepared via a straightforward non-solvent induced phase separation (NIPS) method in most cases A,B as co3) and are typically toxic even at low concentrations [Heavy metals refer to chemical elements that have a high density  membrane in a bid to increase the removal of various heavy metal ions [2 fillers in PVDF matrix, as shown in 2 with the heavy metal ions, the MMM was capable of removing heavy metals to a larger extent (up to a 20% increase in rejection) because of the enhanced adsorption. It was also postulated that the presence of inner-sphere complexes  led to the electrostatic attraction between SnO2 and metal ions which enhanced the adsorption of heavy metals onto the MMM [2/g for UiO-66 [2+ rejection) of MMMs incorporated with MOFs when removing trace amounts of mercury in the feed solution (2.61 ppm) [y metals ,37,42. Ttal ions   which e.61 ppm) . The keyOverall, the various methods to remove heavy metals using MMMs in The adsorption rates might not occur uniformly throughout the filler material for that some functional groups of the material might have more favorable sites for adsorption or higher ion exchange capacities ,39,43,463AsO4 will be transformed into AsO43−) as well as the charge of the membrane surface and heavy metal ions, which ultimately dictates the ion exchange capacity between the MMM and metal ions [4−) and dichromate (Cr2O72−) are the dominant species [It is also worth noting that the pH of the feed is typically adjusted to maximize the adsorption capacity of the MMM for that the former controls the speciation of the heavy metal , thereby freeing up the adsorption sites  or Freundlich (multi-layer adsorption) isotherms ,44, bothon sites D. In anoSize exclusion and Donnan exclusion effects. Besides adsorption, another facile yet promising method to increase the heavy metal removal efficiency of membranes is via size exclusion or Donnan exclusion effects. The latter has shown success by introducing functionalized adsorbents that are of opposite charge to the target metal ions [tal ions . Becausetal ions ,41,43. Ftal ions . In anottal ions . It was 2/g for alumina particles [2O3 nanoparticles) can lead to agglomeration, which is undesirable because it minimizes the surface area for adsorption. Hence, it is essential to modify them . Alsarticles  and 50 narticles ) to maxiarticles ) to reduDyes are colored substances that establish chemical bonds to the substrates (which they are applied onto) such as paper, leather, and textiles. It has been estimated that wastewater containing textile dyes contributes to ~18% of industrial water pollution  and thatAdsorption of dye molecules. A common approach to increase the dye rejection of membranes is to incorporate adsorbent filler materials into the membrane matrix (2 [e matrix . For exaatrix (2 ) into thatrix , and decreased fouling propensity (organic and biofouling), all of which is attributed to the presence of filler material ,55. It ion rates . The keyon rates . The keyPhoto-degradation of dye molecules. It was reported that MMMs containing filler material MIL-125 (Ti) (+) and superoxide free radicals are produced [•OH radicals are known to destroy the cell walls of bacteria).125 (Ti) B were ab125 (Ti) C. First,produced . The proDonnan exclusion and size effects. The optimization of membrane surface charge or pore size is another facile way to increase the dye rejection of porous membranes. For example, the incorporation of filler materials to render the membrane surface more negatively charged has shown success in increasing dye rejection (for negatively charged dyes such as acid black) because of electrostatic repulsion effects [ effects ,53,55. S effects ,53. The  effects . However effects . Hence iPolar organic compounds typically partition into the aqueous phase, thereby exhibiting high mobility in water bodies and within the hydrological cycle. Recent evidence has suggested that trace concentrations of dissolved organic pollutants in water can disrupt the food chain in aquatic ecosystems and the biological functions of living organisms . One of Sieving and electrostatic repulsion effects in dense membranes. The transport phenomena in the dense selective layer of reverse osmosis (RO) membranes are well described by the solution-diffusion mechanism whereby water and solutes dissolve onto the membrane surface and thereafter diffuse through a selective layer. To increase the rejection of organic compounds in dense membranes, a potential approach is to introduce filler materials to modify the charge and pore properties of the polyamide layer. Albergamo et al. explored the use of MMM-RO membranes incorporated with zeolites and aquaporins to treat brackish water loaded with 30 persistent organic micropollutants  [ diuron) . It was  diuron) ). It was diuron) . This su diuron) ) might hIncrease hydrophilicity and adsorption capacity in porous membranes. To increase the retention of OPs in porous membranes  [2 fits the bill of an adsorbent material because of its uniform distribution of mesopores (pore diameter of 3.58 nm), high surface area (350 m2/g), and most importantly, it is mechanically stable [2. To maximize the adsorption capacity of the MMM, it is crucial to disperse the filler materials in the UF membrane matrix to ensure all the pore cavities are available for physisorption.embranes A, the tys (PAHs) . Being ay stable . The mecDual-function MMMs have also been explored, whereby two filler materials are incorporated into the UF membrane matrix to harness the synergistic advantages of both fillers. For example, Anjum et al. reported the success of dual-function UF-MMMs incorporated with UiO-66 and Zeolite 4A for flux enhancement (~2 times higher) at a fairly similar humic acid rejection when compared against the control membrane . This wo3 and NH4+, respectively [3) is toxic to aquatic ecosystems, and its concentration is typically low (<0.05 mg/L) due to the natural replacement of water with freshwater, as well as the biological conversion of ammonia to nitrate in biological processes . However, this process is slow due to the long start-up time and is not effective when the concentration of ammonia nitrogen is high. The excessive presence of nitrate in water bodies will also alter ecological balances due to the growth of algae which consequently inhibits the healthy growth of aquatic plants and animals. To reduce the concentration of ammonia in water bodies, various methods can be used, such as stripping using hydrophobic membrane contactors or nitrification-denitrification process using micro-organisms [Ammonia nitrogen and nitrates are present in the environment due to the excretion of fish waste in water bodies. In aquatic chemistry, both the molecular and ionic forms of ammonia nitrogen exist in the form of NHectively . The molrganisms . However2O3) [4+ and NO3− ions can be readily adsorbed onto zeolite and hematite particles, respectively, because of electrostatic attractions [+, Mg2+, which are commonly found in wastewater) will affect the adsorption capacity of the MMM for that the former can compete with NH4+ for the adsorption sites [Membrane processes using porous UF membranes have been explored to remove ammonia and nitrates in aqueous systems . In rece2O3)  nanopartractions ,35. Howeon sites  is needeEmpirical evidence suggests that the adsorption capacity of the MMM is dictated by the thermodynamic equilibrium process whereby the MMM’s maximum adsorption capacity is dictated by the amount of ions adsorbed as well as the amount of ions in the feed water . The regProcess optimization to improve removal efficiency. One of the primary concerns of membranologists developing novel membranes and membrane processes is how to balance the trade-off between membrane permeability and selectivity [ectivity ,65. To mectivity ,67,68. TProcess combination to improve removal efficiency. Pollutant removal processes using MMMs can be combined with other membrane processes for further improvement of removal efficiency. An RO process is a typical example that can be used in series to post-treat the effluent produced from UF using MMMs. An RO membrane consists of a polyamide active layer that can separate organic and ionic species by providing much dense and charged channels. However, the much dense and charged active layer of RO membranes could be a double-edged sword in that it can lead to the improved removal efficiency of the final product at the expense of lower overall productivity (in terms of water permeance) because of the permselectivity trade-off. Fortunately, the trade-off can be partially relieved by support modification [fication ,70,71, wA dynamic membrane separation (DMS) process  could beNanomaterials have been widely used for pollutant removal as additives to prepare MMMs or building blocks for laminate membranes. Between the two approaches, nanomaterials could find a sustainable and feasible way for pollutant removal through the incorporation into MMMs instead of being used for laminate membranes, as discussed earlier in Diverse kinds of biomasses are listed in Biomass can be converted into a wide variety of carbon-based materials such as biochar, carbon black, activated carbon, carbon nanotubes (CNTs), and GO via carbonization at 600–1200 °C as described in Million tons of non-biodegradable plastic wastes have been annually discarded in the environment and weathered for the last decades, ending up with microplastics (MPs) . MPs areEmpirical evidence has also shown that MPs tend to degrade into smaller fragments  , and tra3·H2O at pH 4.0 [On the other hand, the potential influence of surface characteristics on the interaction between MPs and surfaces could be whittled down when MPs are larger than 10 μm. Instead, the pore system effects including morphology become predominant in the removal of MPs larger than 10 μm . In detat pH 4.0 . This ca2+: 44.4%; Cd2+: 42.8%), demonstrating a need for further improvement in removal efficiency. Moreover, high removal efficiency should be pursued, taking into account that the need for more environmentally friendly materials is growing as many countries have sought to attain a sustainable and green economy to overcome global climate change [This paper covers an overview of the research progresses and trends on the emerging materials used to prepare MMMs for pollutant removal in the recent five years. For the past five years, various types of emerging materials ranging from organic to inorganic nanomaterials have been used to prepare high-performance MMMs for pollutant removal, and their effectiveness has been investigated using a range of pollutants encompassing heavy metals, dye molecules, humic acid, organic compounds, nitrates, ammonia, and so on. Most of the MMMs prepared with novel additives successfully removed more than 90% of pollutants in an aqueous phase. However, some contaminants exhibited lower removal efficiency than 50% (e.g., Nie change ,90. From


Arising from Rohr J. J. et al. npj Microgravity 10.1038/s41526-020-00119-3 (2020)1 that was published in npj Microgravity aimed to provide a quantitative analysis of optic nerve and optic nerve sheath (ONS) cross-sectional areas, and optic nerve deviation, an indication of tortuosity, before and after long-duration spaceflight (LDSF). High-resolution, T1-weighted sagittal, and T2-weighted coronal magnetic resonance imaging (MRI) scans were collected from ten astronauts undergoing ∼6-month missions on the International Space Station (ISS). Each astronaut underwent MRI scanning before flying to the ISS and at five recovery time points after return to Earth . On average, optic nerve cross-sectional areas tended to decrease in their cohort immediately post flight. However, only the 30 days postflight time point showed a statistical difference from preflight, with an average reduction in optic nerve cross-sectional area of −0.89 mm2. The exact reason for this optic nerve area reduction is unclear. As noted by the authors, although increased cerebrospinal fluid (CSF) pressure surrounding the optic nerve can lead to axoplasmic flow stasis and resultant optic disc edema, it is unclear how these changes may specifically impact the optic nerve morphology during and after LDSF. They also documented no significant increase in the ONS diameter following LDSF except in one astronaut with unilateral grade 1 optic disc edema. This suggests that, in nine of the ten astronauts, intracranial pressure was not pathologically elevated immediately after spaceflight. The authors appropriately mention that the ONS in astronauts may demonstrate hysteresis such that exposure to ONS pressure beyond a certain point may result in failure of the sheath to regress to normal following spaceflight and change its response to subsequent pressure changes. It would be interesting to know how many of the ten astronauts they report had previous spaceflights prior to their MRI study that might have impacted their ONS response to repeat extended microgravity exposure.The recent study by Rohr et al.1. They discuss a retrospective MR study by Kramer et al. published in 20122. In that study, the authors observed a central area of T2 hyperintensity in the optic nerve in 26 of the 27 astronauts (96%) with a history of microgravity exposure2. As discussed below, we propose that this T2 hyperintensity in the optic nerve reflects a glymphatic pathway and that altered dynamics of the optic nerve glymphatic circulation may contribute to the reduction in the optic nerve cross-sectional area observed after spaceflight.The authors suggest that a possible explanation for the decrease in optic nerve cross-sectional area could be the role of changes in the optic nerve vasculature2 already reflected signs of the existence of glymphatic flow of CSF into the optic nerve. The “glymphatic system” is a recently discovered brain-wide clearance system in which CSF enters the brain via periarterial channels and exchanges with interstitial fluid3. This fluid is then removed from the brain along perivenous pathways and eventually cleared via the cervical lymphatic vasculature3. CSF within the subarachnoid space (SAS) is gradually forced into the Virchow-Robin spaces by the combined effects of arterial pulsatility, respiration, slow vasomotion, and CSF pressure gradients5. Thereafter, CSF passes into the dense brain parenchyma via perivascular astrocytic aquaporin-4 (AQP4) water channels4.We raise the question of whether the previously reported T2 hyperintensity in the optic nerve of astronauts6. Recently, Mathieu et al.6 provided evidence to support the existence of a glymphatic pathway in the optic nerve following tracer injection into the CSF of live mice. Their findings indicated that CSF enters the optic nerve via spaces surrounding blood vessels, bordered by AQP4-positive astrocytic endfeet. We recently hypothesized that the forcing of perioptic CSF into the perivascular spaces surrounding the central retinal artery, as a result of long-standing microgravity fluid shifts, may partly explain the optic disc edema observed in astronauts during LDSF7. It is important to note that individual variations in ONS anatomy and compliance may impact the end result of this process7. The MR study of astronauts by Kramer et al.2 seems to support the possibility that the optic nerve communicates directly with its SAS. In cases with an ONS kink, the authors documented a significant increase in the diameter of the central area of T2 hyperintensity in the mid optic nerve2. They also noted that the kink produced an increase in ONS diameter suggestive of a localized rise in CSF pressure. The authors hypothesized that a kink may distort the complex system of arachnoid trabeculae, pillars, and septa within the perineural SAS2. This ONS deformation could lead to a restriction of anterograde CSF flow and a resultant pressure gradient2. Furthermore, this process could result in the proximal congestion of the perivascular space of the optic nerve via communication with the perineural SAS2. In another study by Riascos et al.8 of 21 astronauts, MR examination documented a central T2 hypointensity in the epicenter of the previously described T2 hyperintensity in the optic nerve in all those examined, including the two astronauts not yet exposed to microgravity, suggesting the T2 hypointensity may have been created by a central retinal vessel flow void artifact.The optic nerve, a white matter tract of the CNS consisting of afferent axons of the retinal ganglion cells, is also surrounded by CSF within the orbital SAS7, we hypothesize that in those astronauts without pathologically elevated postflight CSF pressures within the ONS, the rapid fluid redistribution upon return to Earth, which consequently may lead to reduced CSF volume and pressure within the ONS, may decrease CSF inflow along the optic nerve periarterial glymphatic spaces. As a result, after landing on Earth, these periarterial spaces may dramatically reduce and even disappear, at least partly contributing to the observed decrease in optic nerve cross-sectional area. In order to further explore and validate this hypothesis, it would be interesting to measure changes in the diameter of the central area of T2 hyperintensity in the optic nerve before, as soon as possible after, and at several more recovery time points following spaceflight.Considering the above, we believe that the reduction in optic nerve cross-sectional area observed after spaceflight may result, at least partly, from altered dynamics of the optic nerve glymphatic circulation. Indeed, while prolonged exposure to microgravity may predispose to an overload in the periarterial inflow of CSF into the optic nerve


Tumour necrosis factor (TNF) is an inflammatory cytokine produced in response to viral infections that promotes the recruitment and activation of leukocytes to sites of infection. This TNF-based host response is essential to limit virus spreading, thus poxviruses have evolutionarily adopted diverse molecular mechanisms to counteract TNF antiviral action. These include the expression of poxvirus-encoded soluble receptors or proteins able to bind and neutralize TNF and other members of the TNF ligand superfamily, acting as decoy receptors. This article reviews in detail the various TNF decoy receptors identified to date in the genomes from different poxvirus species, with a special focus on their impact on poxvirus pathogenesis and their potential use as therapeutic molecules. TNF is a potent pro-inflammatory cytokine with a broad range of biological effects, ranging from the activation of inflammatory gene programs to cell differentiation or apoptosis induction while also playing an essential role in the host control of many viral infections ,2. It waTNF is first produced as a 26 kDa class II transmembrane protein (tmTNF) on the surface of cells. Then, tmTNF is cleaved by the metalloprotease TNF-converting enzyme (TACE) to be released as a soluble homotrimeric cytokine (sTNF) of about 51 kDa that can execute its function far away from the site where it is synthesized [Far from just being a precursor, tmTNF is a key regulator for many immune and inflammatory processes, as both forms of TNF (tmTNF and sTNF) can bind to two different transmembrane cellular receptors with different affinities . WhereasSoluble versions of TNFR1 and TNFR2 may also exist as a result of alternative splicing or proteolytic cleavage ,11. To rIn addition, tmTNF can induce reverse signalling, a process by which this membrane-bound cytokine can also act as a receptor and trigger intracellular signalling in tmTNF-bearing cells. This tmTNF-elicited reverse signalling is key for the activation and regulation of T cells, macrophages and natural killer (NK) cells ,16.Another TNFLSF member is lymphotoxin-α (LTα), the closest homolog to TNF with 51% sequence similarity, which mainly exists as a soluble homotrimer secreted from T and B lymphocytes. LTα differs from TNF in the amino terminal part, which resembles a typical signal peptide, making LTα secretion extremely efficient. Indeed, unlike most members of TNFLSF, LTα can only anchor to the cell surface after association with membrane-bound lymphotoxin-β (LTβ) to form a heterotrimer . LTα binOne of the main physiological roles of TNF is the mediation of acute inflammation during virus infection. TNF is locally induced early after virus infection and enhances its own expression together with other pro-inflammatory cytokines and also the secretion of chemokines that promote the recruitment and activation of leukocytes. In animal models of infection, TNF, synergistically with interferon (IFN) γ, has been shown to interfere with the replication of diverse viruses, such as hepatitis B virus  or murinThe specific relevance of TNF and TNF signalling in the protection against poxvirus infections is also largely known and supported by experimental data from in vivo studies. For instance, mouse strains naturally resistant to a lethal poxvirus infection, such as ectromelia virus (ECTV), became more susceptible after TNFR deletion, since 60% of the knockout animals succumbed to infection . Indeed,Poxviruses are known for their broad repertoire of immunomodulatory proteins dedicated to evading the host response during infection. One of their most distinctive immune evasion mechanisms is the secretion of viral proteins from infected cells with the ability to bind key host cytokines before engagement with cellular receptors, thus acting as decoy receptors . These pvTNFRs are soluble proteins that mimicking the extracellular domains of TNFRSF and bind to members of TNFLSF in solution, preventing their interaction with cognate receptors at the cell surface . To dateAlthough the structure of just one vTNFR has been experimentally determined , it is wAs stated before, vTNFRs represent soluble viral versions of the membrane-anchored cellular receptors. However, VACV CrmE can additionally associate to the cell surface, maintaining its TNF binding properties intact. Since CrmE lacks a transmembrane domain, this association might occur through interaction with cellular proteins, remaining attached to the cell surface before its cleavage to act as a soluble inhibitor. Alternatively, CrmE might be retained through its N-terminal region at the cell surface, allowing CRDs exposure to bind soluble TNF .Diversity also extends to the number of gene copies encoding vTNFRs among different poxviruses. For example, while one copy of the CrmC gene is found in CPXV, CrmB and CrmD are encoded by two gene copies in CPXV and ECTV, respectively. The number of gene copies encoding the same vTNFR can even vary across poxvirus species to influence its expression levels. Remarkably, although CrmD is expressed at lower levels than CrmC by CPXV, two gene copies in ECTV provide higher expression levels of CrmD during infection .One of the most fascinating aspects of vTNFRs is their degree of specificity to bind diverse members of the TNFLSF together with their specialization to discriminate them from different animal species. CrmC  and CrmEIt is worth noting that vTNFRs CrmB, C, D and E are differentially distributed among poxvirus species. It seems that those poxviruses with a strict host range, like variola virus (VARV) and ECTV, usually express just one of them. For instance, a strictly human poxvirus like VARV only expresses CrmB as functional vTNFR and the mouse-specific ECTV expresses only CrmD ,62,63. OThis specific disposal of vTNFRs among viral species seems to be the result of their co-evolution with their hosts and would indicate that poxviruses have adapted their anti-TNF strategies to the specific TNF response they need to counteract. For example, although CrmB orthologues encoded by CPXV, MPXV, and VARV show high sequence similarity, they exhibit very different ligand binding properties. CrmB from VARV is the most effective at binding and inhibiting human TNF . SimilarIt is also remarkable that CrmC and CrmE can bind both human and mouse TNF with high affinity, but they are only able to inhibit the biological activity of mouse TNF and human TNF, respectively. This may be explained by specific differences in how vTNFRs recognize their ligands, as binding nonessential residues may increase affinity but may not interfere with the cellular recognition, thus being irrelevant for neutralizing activity ,52. RegavTNFRs are considered more similar to TNFR2, as they share more sequence similarity with the extracellular domain of TNFR2 (32%) than to the one of TNFR1 (22%). In addition, the structure of CrmE , the uniMost of the tmTNF activity overlaps with sTNF, however tmTNF can additionally trigger reverse signalling after interaction with TNFR at the cell surface and dampen inflammatory processes ,38,40,72The particular, the behaviour of CrmE deserves a special mention, as it binds both murine sTNF and tmTNF with high affinity but can only inhibit murine tmTNF activity. These data suggest that the molecular structures from mouse tmTNF and sTNF are different and raise two possibilities to explain this surprising behaviour: either tmTNF and sTNF bind to CrmE through different residues, or these interacting residues are the same, but the biological activity of these cytokines relies on different sites. In any case, the contribution of tmTNF signalling to the defence against poxvirus infections in mice becomes evident due to the fact that the expression of recombinant CrmE surprisingly enhanced the virulence of a VACV strain lacking other vTNFRs . Besides their CRDs, CrmB and CrmD contain an additional C-terminal domain, termed Smallpox virus-Encoded Chemokine Receptor (SECRET), with the ability to bind with high affinity and block the activity of a particular set of chemokines involved in mucosal and skin inflammation. This domain shares no sequence similarity to cellular TNFRs or any other known cellular protein and represents an independent folding domain that can exert its function either independently or fused to a TNFR . The modAlthough the SECRET domain is not related to any cellular gene, sequence analysis of poxviral genomes revealed three additional genes encoding separate SECRET domain-containing proteins (SCPs) . DespiteA fine characterization of the functional boundaries between CRDs and the SECRET domain from CrmD has recently confirmed that CRD4, although initially assigned to the TNF binding region, contributes to the SECRET domain-mediated chemokine binding. This incertitude was already hinted at when Asp167 and Glu169 residues in CRD4 from CrmD were identified as key chemokine-binding determinants in the structure of the SECRET domain–CX3CL1 complex . In lineT2 protein was the first vTNFR identified in poxvirus  and to dGiven its dual activity as a TNF and apoptosis inhibitor, it is not surprising that M-T2 was addressed as an obvious virulence factor. Indeed, the infection of susceptible European rabbits with MYXV lacking both gene copies encoding M-T2 resulted in disease attenuation and diminished mortality, indicating a critical role for this vTNFR in the development of myxomatosis .vCD30 was identified as a soluble viral version of the cellular receptor CD30, which is also known as TNFRSF8 and expressed by a small subset of activated T and B cells . vCD30 iIn addition to vTNFRs, some poxvirus species have developed their own vTNFBPs to impede TNF signalling. The 2L protein was the first vTNFBP, with no sequence homology to any known cellular TNFR, identified from Tanapox virus (TPXV)-infected cell supernatants. Later, two homologs were found in the other two existing yatapoxvirus: Yaba-like disease virus (YLDV) and Yaba monkey tumor virus (YMTV). Similar to some vTNFRs, 2L binds human TNF with very high affinity to prevent cytokine signalling through TNFR1 and TNFR2 , howeverThis structural analysis also showed that the 2L interaction with TNF resembles the TNF interaction with the cellular TNFR, a central TNF trimmer bound by three symmetrically arranged 2L molecules. These data raise the hypothesis that 2L might have originated from a fragmented gene encoding an MHC-I molecule that was subsequently incorporated and modified by poxviruses to bind and block TNF.Besides the 2L protein from yatapoxviruses, proteins SPV003 and DPV008 from swinepox virus and deerpox virus, respectively, are also considered as vTNFBPs ,92. In cAlthough we have previously mentioned the relative importance of CD30 and T2, the contribution of vTNFRs to poxvirus pathogenesis was initially unclear, as most reports generating viruses lacking expression of vTNFRs showed very limited effects or were not conclusive. For example, deletion of CrmB resulted in an attenuated phenotype compared to parental virus in an intracranial model of CPXV , a routeIn contrast, CrmD is the only functional TNF inhibitor in ECTV, a strict mouse pathogen causing mousepox disease, making ECTV infection of susceptible mice a suitable model to study the contribution of this vTNFR to poxvirus pathogenesis. In contrast with the observations described above, the infection of susceptible BALB/c mice with an ECTV lacking CrmD expression (ECTV∆CrmD), and therefore in the absence of any TNF blocking activity, resulted in a dramatic virus attenuation and a difference of six orders of magnitude in LD50 compared to wild type virus . In the TNF modulation by ECTV represents an excellent example illustrating how virus evasion strategies can increase the understanding of the host immune regulation mechanisms. In this sense, the analysis of the specific contribution to mousepox pathogenesis of CRDs and SECRET domains from CrmD revealed that chemokines and TNF cooperate during virus infection to develop the inflammatory response and provide anti-viral immunity. Thus, the reinsertion of either the anti-TNF (CRDs domain) or the anti-chemokine (SECRET domain) activities into ECTV∆CrmD did not restore full virulence, indicating that both functions need to operate together to evade the potent antiviral response. In line with this, a recent study evidenced the deleterious consequences of an excessive TNF based response during poxvirus infections. The intranasal infection of mouse strains genetically resistant to mousepox with ECTV∆CrmD caused uniform mortality due to excessive TNF action and a dysregulated inflammatory response . In thisDysregulated TNF production has also been postulated as being responsible for the pathology in many inflammatory disorders, such as rheumatoid arthritis, psoriasis, vasculitis or Crohn's disease . In thesSome vTNFRs show broader ligand specificity or are more potent inhibitors of TNF signalling than etanercept , supportChemokines are also considered an important target for the development of anti-inflammatory therapies, as they mediate leukocyte migration . As descCrmD has also inspired an additional modification related to ligand specificity intended to prevent some of the adverse effects from actual anti-TNF therapies . EtanercvTNFRs could have an additional application in poxvirus vaccination. Although current vaccines against virulent poxviruses have proven their efficacy in the past, there is still uncertainty about the duration of their immunity and their adverse effects, raising considerable interest to develop new and safer vaccines. In this sense, immunization with CrmD protects mice from otherwise lethal mousepox, most likely due to antibody-mediated blockade of CrmD . A similIn conclusion, these soluble decoy receptors constitute a highly efficient strategy to prevent TNF effects, as deduced from the recent evidence demonstrating the relevant contribution of vTNFRs to poxvirus virulence and pathogenesis. Thus, further research on this repertoire of TNF decoy receptors might inspire additional modifications to improve current therapies for inflammatory diseases, or even the generation of new anti-inflammatory molecules with a viral origin.