miR-200c-3p regulates α4 integrin-mediated T cell adhesion and migration.

A microRNA miR-200c-3p is a regulator of epithelial-mesenchymal transition to control adhesion and migration of epithelial and mesenchymal cells. However, little is known about whether miR-200c-3p affects lymphocyte adhesion and migration mediated by integrins. Using TK-1 (a T lymphoblast cell) as a model of T cell, here we show that repressed expression of miR-200c-3p upregulated α4 integrin-mediated adhesion to and migration across mucosal addressin cell adhesion molecule-1 (MAdCAM-1). Conversely, overexpression of miR-200c-3p downregulated α4 integrin-mediated adhesion and migration. Unlike in epithelial cells, miR-200c-3p did not target talin, a conformation activator of integrin, but, targeted E26-transformation-specific sequence 1 (ETS1), a transcriptional activator of α4 integrin, in T cells. Treatment of the miR-200c-3p-low-expressing TK-1 cells that possessed elevated α4 integrin with ETS1 small interfering RNA (siRNA) resulted in the reversion of the α4 integrin expression, supporting that ETS1 is a target of miR-200c-3p. A potential proinflammatory immune-modulator retinoic acid (RA) treatment of TK-1 cells elicited a significant reduction of miR-200c-3p and simultaneously a marked increase in ETS1 and α4 integrin expression. An anti-inflammatory cytokine TGF-ß1 treatment elevated miR-200c-3p, thereby downregulating ETS1 and α4 integrin expression. These results suggest that miR-200c-3p is an important regulator of α4 integrin expression and functions and may be controlled by RA and TGF-ß1 in an opposite way. Overexpression of miR-200c-3p could be a novel therapeutic option for treatment of gut inflammation through suppressing α4 integrin-mediated T cell migration.

Overexpression of putative glutathione peroxidase from Neopyropia-associated microorganisms in Chlamydomonas to respond to abiotic stress.

Lipid accumulation in microalgae can be substantially enhanced by exposing the microalgae to abiotic stress, thus increasing biofuel production. However, this also generates reactive oxygen species (ROS), which disrupts cell metabolism and reduces their productivity. Previous mRNA sequencing analyses in Neopyropia yezoensis and its associated microorganisms elucidated a putative glutathione peroxidase (PuGPx) gene. Here, this putative glutathione peroxidase was overexpressed in the microalga Chlamydomonas reinhardtii, which increased cell growth and survival rates compared to the control group under abiotic stress. Additionally, increased lipid accumulation was observed under salinity stress, high-temperature stress, and hydrogen peroxide (H2O2)-induced oxidative stress. These results suggest that PuGPx plays a protective role against abiotic stress in C. reinhardtii and stimulates lipid accumulation, which could be considered advantageous in terms of biofuel production.

Recombinant soluble thrombomodulin attenuates cisplatin-induced intestinal injury by inhibiting intestinal epithelial cell-derived cytokine secretion.

BACKGROUND: Intestinal injury is one of the main side-effects of cisplatin chemotherapy, impairing the quality of life in patients with cancer. In this study, we investigated the protective effects of recombinant soluble thrombomodulin (rsTM), which is a potent anti-inflammatory agent, on cisplatin-induced intestinal injury. METHODS: We first evaluated the effects of rsTM on intestinal injury caused by cisplatin in mice in vivo. Disease progression was monitored by analyzing loss of body weight and histological changes in intestinal tissue. We then investigated the effects of rsTM on mouse intestinal organoid formation and growth in vitro. Gene expression levels were analyzed by quantitative real-time polymerase chain reaction and Western blotting. RESULTS: rsTM treatment significantly attenuated the loss of body weight, histological damage and gene expression levels of pro-inflammatory cytokines such as interleukin-6, tumor necrosis factor-α and high-mobility group box-1 in a cisplatin-treated mouse model. Furthermore, rsTM alleviated the inflammatory response and apoptosis in a cisplatin-treated intestinal epithelial organoid model. CONCLUSION: rsTM suppresses cisplatin-induced intestinal epithelial cell-derived cytokine production and alleviates intestinal mucositis.

Vitamin D deficiency and mortality among critically ill surgical patients in an urban Korean hospital.

Critically ill patients in intensive care units (ICUs) are exposed to various risk factors for vitamin D deficiency. Vitamin D deficiency in extended-stay patients may result in decreased muscle mass and increased fat tissue, which may impair rehabilitation and recovery. Our study aimed to evaluate the degree of serum vitamin D deficiency in critically ill surgical patients and its association with clinical outcomes. Clinical data from 186 adult male (n = 121; 65.1%) and female (n = 65; 34.9%) patients hospitalized in surgical ICUs at Ajou University Hospital from April 2015 to September 2016 were retrospectively analyzed. All adult surgical patients between the age of 18 and 88 years were enrolled. The mean serum 25-hydroxyvitamin D (25[OH]D) level of all patients was 17.8 ng/mL. A total of 120 patients (64.5%) with serum 25(OH)D levels < 20 ng/mL were classified as the deficiency group. A prolonged hospital stay was observed among the deficiency group but was not statistically significant (p = 0.824). Serum 25(OH)D levels were significantly correlated with age but inversely correlated with Sequential Organ Failure Assessment (SOFA) score, selenium, triglycerides, and C-reactive protein levels. There was no significant difference in mortality rates between the group with a vitamin D injection and the group without a vitamin D injection (14.6% vs. 16.9%, p = 0.074). Vitamin D deficiency was common in surgical ICU patients; however, vitamin D levels were higher in older patients. In conclusion, vitamin D deficiency was inversely associated with the SOFA severity score (correlation coefficient -0.165, p = 0.024) but was not associated with the length of hospital or ICU stay and mortality.

Recombinant soluble thrombomodulin accelerates epithelial stem cell proliferation in mouse intestinal organoids and promotes the mucosal healing in colitis.

BACKGROUND AND AIM: Epithelial regeneration, a critical step for the mucosal healing in inflammatory bowel disease, is tightly regulated by stem cells. Therefore, identification of the specific factors that induce stem cell proliferation could contribute to the development of effective strategies for treating inflammatory bowel disease. Recombinant soluble thrombomodulin (rsTM) has previously been shown to promote cell proliferation in skin and corneal wound healing in murine models, but its effects on intestinal epithelial cell proliferation remains unclear. METHODS: Mouse intestinal organoids and dextran sulfate sodium (DSS)-induced colitis mouse model were used to assess the effects of rsTM on proliferation of intestinal epithelial cells. The size and budding morphologies of organoids were studied by confocal microscopy. The gene expression levels were analyzed by quantitative real-time polymerase chain reaction and immunofluorescence analysis. The effects of rsTM on DSS-induced colitis were investigated by evaluating body weight changes, colon length, histological score, and survival rate. RESULTS: The rsTM markedly stimulated the growth of intestinal organoids, thereby increasing the surface areas and budding phenotypes of the organoids. rsTM also significantly upregulated the gene expression of intestinal stem cell-specific and epithelial cell-specific markers in a dose-dependent manner. Furthermore, the treatment with high concentrations of rsTM significantly improved the recovery of body weight, histological outcomes, colon length shortening, and prolonged the survival of mice with colitis. CONCLUSIONS: The rsTM promotes intestinal stem cell proliferation in intestinal organoids and enhances the mucosal healing during recovery phase in DSS-induced colitis.

Integrin Regulation in Immunological and Cancerous Cells and Exosomes.

Integrins represent the biologically and medically significant family of cell adhesion molecules that govern a wide range of normal physiology. The activities of integrins in cells are dynamically controlled via activation-dependent conformational changes regulated by the balance of intracellular activators, such as talin and kindlin, and inactivators, such as Shank-associated RH domain interactor (SHARPIN) and integrin cytoplasmic domain-associated protein 1 (ICAP-1). The activities of integrins are alternatively controlled by homotypic lateral association with themselves to induce integrin clustering and/or by heterotypic lateral engagement with tetraspanin and syndecan in the same cells to modulate integrin adhesiveness. It has recently emerged that integrins are expressed not only in cells but also in exosomes, important entities of extracellular vesicles secreted from cells. Exosomal integrins have received considerable attention in recent years, and they are clearly involved in determining the tissue distribution of exosomes, forming premetastatic niches, supporting internalization of exosomes by target cells and mediating exosome-mediated transfer of the membrane proteins and associated kinases to target cells. A growing body of evidence shows that tumor and immune cell exosomes have the ability to alter endothelial characteristics (proliferation, migration) and gene expression, some of these effects being facilitated by vesicle-bound integrins. As endothelial metabolism is now thought to play a key role in tumor angiogenesis, we also discuss how tumor cells and their exosomes pleiotropically modulate endothelial functions in the tumor microenvironment.

Remodeling of Bone Marrow Niches and Roles of Exosomes in Leukemia.

Leukemia is a hematological malignancy that originates from hematopoietic stem cells in the bone marrow. Significant progress has made in understanding its pathogensis and in establishing chemotherapy and hematopoietic stem cell transplantation therapy (HSCT). However, while the successive development of new therapies, such as molecular-targeted therapy and immunotherapy, have resulted in remarkable advances, the fact remains that some patients still cannot be saved, and resistance to treatment and relapse are still problems that need to be solved in leukemia patients. The bone marrow (BM) niche is a microenvironment that includes hematopoietic stem cells and their supporting cells. Leukemia cells interact with bone marrow niches and modulate them, not only inducing molecular and functional changes but also switching to niches favored by leukemia cells. The latter are closely associated with leukemia progression, suppression of normal hematopoiesis, and chemotherapy resistance, which is precisely the area of ongoing study. Exosomes play an important role in cell-to-cell communication, not only with cells in close proximity but also with those more distant due to the nature of exosomal circulation via body fluids. In leukemia, exosomes play important roles in leukemogenesis, disease progression, and organ invasion, and their usefulness in the diagnosis and treatment of leukemia has recently been reported. The interaction between leukemia cell-derived exosomes and the BM microenvironment has received particular attention. Their interaction is believed to play a very important role; in addition to their diagnostic value, exosomes could serve as a marker for monitoring treatment efficacy and as an aid in overcoming drug resistance, among the many problems in leukemia patients that have yet to be overcome. In this paper, we will review bone marrow niches in leukemia, findings on leukemia-derived exosomes, and exosome-induced changes in bone marrow niches.

Cellular and Exosomal Regulations of Sepsis-Induced Metabolic Alterations.

Sepsis is a sustained systemic inflammatory condition involving multiple organ failures caused by dysregulated immune response to infections. Sepsis induces substantial changes in energy demands at the cellular level leading to metabolic reprogramming in immune cells and stromal cells. Although sepsis-associated organ dysfunction and mortality have been partly attributed to the initial acute hyperinflammation and immunosuppression precipitated by a dysfunction in innate and adaptive immune responses, the late mortality due to metabolic dysfunction and immune paralysis currently represent the major problem in clinics. It is becoming increasingly recognized that intertissue and/or intercellular metabolic crosstalk via endocrine factors modulates maintenance of homeostasis, and pathological events in sepsis and other inflammatory diseases. Exosomes have emerged as a novel means of intercellular communication in the regulation of cellular metabolism, owing to their capacity to transfer bioactive payloads such as proteins, lipids, and nucleic acids to their target cells. Recent evidence demonstrates transfer of intact metabolic intermediates from cancer-associated fibroblasts via exosomes to modify metabolic signaling in recipient cells and promote cancer progression. Here, we review the metabolic regulation of endothelial cells and immune cells in sepsis and highlight the role of exosomes as mediators of cellular metabolic signaling in sepsis.

Distinct Age-Specific miRegulome Profiling of Isolated Small and Large Intestinal Epithelial Cells in Mice.

The intestinal epithelium serves as a dynamic barrier to protect the host tissue from exposure to a myriad of inflammatory stimuli in the luminal environment. Intestinal epithelial cells (IECs) encompass differentiated and specialized cell types that are equipped with regulatory genes, which allow for sensing of the luminal environment. Potential inflammatory cues can instruct IECs to undergo a diverse set of phenotypic alterations. Aging is a primary risk factor for a variety of diseases; it is now well-documented that aging itself reduces the barrier function and turnover of the intestinal epithelium, resulting in pathogen translocation and immune priming with increased systemic inflammation. In this study, we aimed to provide an effective epigenetic and regulatory outlook that examines age-associated alterations in the intestines through the profiling of microRNAs (miRNAs) on isolated mouse IECs. Our microarray analysis revealed that with aging, there is dysregulation of distinct clusters of miRNAs that was present to a greater degree in small IECs (22 miRNAs) compared to large IECs (three miRNAs). Further, miRNA-mRNA interaction network and pathway analyses indicated that aging differentially regulates key pathways between small IECs (e.g., toll-like receptor-related cascades) and large IECs (e.g., cell cycle, Notch signaling and small ubiquitin-related modifier pathway). Taken together, current findings suggest novel gene regulation pathways by epithelial miRNAs in aging within the gastrointestinal tissues.

Intestinal Epithelium-Derived Luminally Released Extracellular Vesicles in Sepsis Exhibit the Ability to Suppress TNF-a and IL-17A Expression in Mucosal Inflammation.

Sepsis is a systemic inflammatory disorder induced by a dysregulated immune response to infection resulting in dysfunction of multiple critical organs, including the intestines. Previous studies have reported contrasting results regarding the abilities of exosomes circulating in the blood of sepsis mice and patients to either promote or suppress inflammation. Little is known about how the gut epithelial cell-derived exosomes released in the intestinal luminal space during sepsis affect mucosal inflammation. To study this question, we isolated extracellular vesicles (EVs) from intestinal lavage of septic mice. The EVs expressed typical exosomal (CD63 and CD9) and epithelial (EpCAM) markers, which were further increased by sepsis. Moreover, septic-EV injection into inflamed gut induced a significant reduction in the messaging of pro-inflammatory cytokines TNF-a and IL-17A. MicroRNA (miRNA) profiling and reverse transcription and quantitative polymerase chain reaction (RT-qPCR) revealed a sepsis-induced exosomal increase in multiple miRNAs, which putatively target TNF-a and IL-17A. These results imply that intestinal epithelial cell (IEC)-derived luminal EVs carry miRNAs that mitigate pro-inflammatory responses. Taken together, our study proposes a novel mechanism by which IEC EVs released during sepsis transfer regulatory miRNAs to cells, possibly contributing to the amelioration of gut inflammation.

Talin-2 regulates integrin functions in exosomes.

Integrins on exosomes have been shown to mediate binding to recipient cells, potentially playing important roles in controlling exosomal internalization and organ distributions. Although the ability of cellular integrins to mediate cell adhesion is known to be regulated by the cytoplasmic adaptor protein talin, whether the activity of exosomal integrins is similarly regulated by talin remains to be elucidated. Here we have studied this question in T-cell exosomes that surface express the integrins αLß2 and α4ß7. T-cells and T-cell exosomes engineered to lack talin-2 showed reduced binding to the integrin ligand ICAM-1 and MAdCAM-1 compared with control T-cells and exosomes, despite the fact that those T cells and exosomes express intact levels of the other isoform talin-1. In addition, talin-2-deficient T-cell exosomes were less efficiently internalized by endothelial cells, compared with control exosomes. These results suggest that the mechanisms of talin-mediated integrin regulation operate similarly in cells and exosomes.

Anti-adhesive effects of human soluble thrombomodulin and its domains.

We reported previously that leukocyte ß2 integrins (LFA-1 and Mac-1) bind to the serine/threonine-rich domain of thrombomodulin (TM) expressed on vascular endothelial cells (VECs). Recombinant human soluble TM (rhsTM, TMD123) has been approved as a therapeutic drug for septic disseminated intravascular coagulation. However, the roles of TMD123 on the adhesion of leukocyte integrins to VECs remain unclear. In the current study, we have revealed that an integrin-dependent binding between human peripheral blood mononuclear cells (PBMCs) and VECs was inhibited by TMD123. Next, using mutant proteins composed of isolated TM extracellular domains, we examined the structural characteristics responsible for the anti-adhesion properties of TMD123. Namely, we investigated whether the effects of the binding of TM and leukocytes was inhibited by the administration of TMD123. In fact, we confirmed that TMD123, TMD1, and TMD3 inhibited the binding of PBMCs to the immobilized recombinant proteins TMD123 and TMD3. These results indicate that TMD123 inhibited the adhesion of leukocytes to endothelial cells via ß2 integrins and endothelial TM. Moreover, since TMD1 might bind to leukocytes via other adhesion receptors than integrins, TMD1 and TMD3 appear to inhibit leukocyte binding to TM on VECs via different mechanisms. In summary, TMD123 (rhsTM), TMD1 or TMD3 is a promising treatment option for sepsis that attenuates integrin-dependent binding of leukocytes to VECs, and may inhibit the undesirable adhesion and migration of leukocytes to VECs in sepsis.

Reduced substrate stiffness promotes M2-like macrophage activation and enhances peroxisome proliferator-activated receptor γ expression.

The increased stiffness of the extracellular microenvironment observed in cancer and atherosclerosis is thought to regulate the activation of tissue-resident immune cells. However, it remains to be determined whether such substrate stiffness affects macrophage activation phenotypes. Here, we have studied the impact of substrate stiffness on in vitro activation phenotypes of the human monocyte cell line THP-1. THP-1 cells were activated while being cultured on 1%, 4%, 10% agarose gel (soft substrate) or on a plastic plate (stiff substrate). We have shown that a soft, versus a stiff, substrate attenuates the pro-inflammatory activity of M1 promoting-activated THP-1 cells. In addition, we have found that M1-related marker expression and phagocytic activity was lower in THP-1 cells activated on a soft substrate compared to cells on stiff substrates. THP-1 cells alternatively activated on soft substrates showed enhanced M2-like phenotypes. We have found that peroxisome proliferator-activated receptor γ (PPARγ) expression was up-regulated in THP-1 cells activated on a soft substrate. We have shown that the PPARγ antagonist GW9662 partially suppresses M2-like activation of THP-1 cells activated on a soft substrate. Substrate stiffness is, therefore, an important factor in regulating the balance of the pro-inflammatory M1 and anti-inflammatory M2 activation phenotypes.

Conditions for laryngeal mask airway placement in terms of oropharyngeal leak pressure: a comparison between blind insertion and laryngoscope-guided insertion.

BACKGROUND: Insertion under laryngoscopic guidance has been used to achieve ideal positioning of the laryngeal mask airway (LMA). However, to date, the efficacy of this technique has been evaluated only using fiberoptic evaluation, and the results have been conflicting. Other reliable tests to evaluate the efficacy of this technique have not been established. Recently, it has been suggested that the accuracy of LMA placement can be determined by clinical signs such as oropharyngeal leak pressure (OPLP). The aim of this study was to assess the efficacy of LMA insertion under laryngoscopic guidance using OPLP as an indicator. METHODS: After approved by the institutional ethics committee, a prospective comparison of 100 patients divided into 2 groups (50 with blind technique and 50 with the laryngoscope technique) were evaluated. An LMA (LarySeal™, Flexicare medical Ltd., UK) was inserted using the blind approach in the blind insertion group and using laryngoscopy in the laryngoscope-guided insertion group. The OPLP, fiberoptic position score, whether the first attempt at LMA insertion was successful, time taken for insertion, ease of LMA insertion, and adverse airway events were recorded. RESULTS: Data were presented as mean ± standard deviation. The OPLP was higher in the laryngoscope-guided insertion group than in the blind insertion group (21.4 ± 8.6 cmH2O vs. 18.1 ± 6.1 cmH2O, p = 0.031). The fiberoptic position score, rate of success in the first attempt, ease of insertion, and pharyngolaryngeal adverse events were similar between both groups. The time taken for insertion of the LMA was significantly longer in the laryngoscope-guided insertion group, compared to blind insertion group (35.9 ± 9.5 s vs. 28.7 ± 9.5 s, p < 0.0001). CONCLUSION: Laryngoscope-guided insertion of LMA improves the airway seal pressure compared to blind insertion. Our result suggests that it may be a useful technique for LMA insertion. TRIAL REGISTRATION: cris.nih.go.kr , identifier: KCT0001945 (2016-06-17).

MicroRNA-mediated dynamic control of mucosal immunity.

The gastrointestinal tract is a complex and important physiological and immunological organ embodying the first line of defense by which mucosal immunity regulates the immense number and diversity of naturally encountered antigens and commensal microflora. Effective microRNA (miRNA) control of transcription factors or mediators in mucosal immunity is essential to host defense and homeostasis in both physiologic and pathologic states. MiRNA biology has advanced our understanding of the immune regulatory system network at the level of post-transcriptional gene modification. Increasing knowledge on circulating miRNAs could potentially enhance diagnostic techniques in inflammatory bowel disease (IBD). Furthermore, recent findings on the dynamic role of exosomes vis-à-vis the intercellular transportation of miRNAs may provide insights on the use of miRNA as a target for treating IBD.

Bone Regeneration using Silk Hydroxyapatite Hybrid Composite in a Rat Alveolar Defect Model.

Background: To overcome the limited source of autogenous bone in bone grafting, many efforts have been made to find bone substitutes. The use of hybrid composites of silk and hydroxyapatite to simulate natural bone tissue can overcome the softness and brittleness of the individual components. Methods: Critical-sized, 7 x 4 x 1.5 mm alveolar defects were created surgically in 36 Sprague-Dawley rats. Three treatment groups were tested: an empty defect group (group I), a silk fibrin scaffold group (group II), and a hydroxyapatite-conjugated silk fibrin scaffold group (group III). New bone formation was assessed using computed tomography and histology at 4, 8, and 12 weeks, and semi-quantitative western blot analysis was done to confirm bone protein formation at 12weeks. Statistical analysis of new bone formation was done using the Kruskal-Wallis test. Results: Radiomorphometric volume analysis revealed that new bone formation was 64.5% in group I, 77.4% in group II, and 84.8% in group III (p=0.027) at 12 weeks. Histologically, the osteoid tissues were surrounded by osteoblasts not only at the border of the bone defect but in the center of the scaffold implanted area in group III from week 8 on. Semi-quantitative western blotting revealed that osteocalcin expression in group III was 1.8 times higher than group II and 2.6 times higher than group I. Conclusions: New bone formation was higher in hybrid scaffolds. Both osteoconduction at the defect margin and osteoinduction at the center of the defect were confirmed. There were no detected complications related to foreign body implantation.

Structural basis of blocking integrin activation and deactivation for anti-inflammation.

Integrins mediate leukocyte accumulation to the sites of inflammation, thereby enhancing their potential as an important therapeutic target for inflammatory disorders. Integrin activation triggered by inflammatory mediators or signaling pathway is a key step to initiate leukocyte migration to inflamed tissues; however, an appropriately regulated integrin deactivation is indispensable for maintaining productive leukocyte migration. While typical integrin antagonists that block integrin activation target the initiation of leukocyte migration, a novel class of experimental compounds has been designed to block integrin deactivation, thereby perturbing the progression of cell migration. Current review discusses the mechanisms by which integrin is activated and subsequently deactivated by focusing on its structure-function relationship.

Comparative metabolism of fargesin in human, dog, monkey, mouse, and rat hepatocytes.

Fargesin, a bioactive lignan derived from Flos Magnoliae, possesses anti-inflammatory, anti-oxidative, anti-melanogenic, and anti-apoptotic effects. This study compared the metabolic profiles of fargesin in human, dog, monkey, mouse, and rat hepatocytes using liquid chromatography-high resolution mass spectrometry. In addition, we investigated the human cytochrome P450 (CYP), UDP-glucuronosyltransferase (UGT), and sulfotransferase (SULT) enzymes responsible for fargesin metabolism. The hepatic extraction ratio of fargesin among the five species ranged from 0.59 to 0.78, suggesting that it undergoes a moderate-to-extensive degree of hepatic metabolism. During metabolism, fargesin generates three phase 1 metabolites, including fargesin catechol (M1) and O-desmethylfargesin (M2 and M3), and 11 phase 2 metabolites, including O-methyl-M1 (M4 and M5) via catechol O-methyltransferase (COMT), glucuronides of M1, M2, M4, and M5, and sulfates of M1-M5. The production of M1 from fargesin via O-demethylenation is catalyzed by CYP2C9, CYP3A4, CYP2C19, and CYP2C8 enzymes, whereas the formation of M2 and M3 (O-desmethylfargesin) is catalyzed by CYP2C9, CYP2B6, CYP2C19, CYP3A4, CYP1A2, and CYP2D6 enzymes. M4 is metabolized to M4 glucuronide by UGT1A3, UGT1A8, UGT1A10, UGT2B15, and UGT2B17 enzymes, whereas M4 sulfate is generated by multiple SULT enzymes. Fargesin is extensively metabolized in human hepatocytes by CYP, COMT, UGT, and SULT enzymes. These findings help to elucidate the pharmacokinetics and drug interactions of fargesin.

Integrin-Mediated Exosomal Homing to Organs.

Exosomes, the small extracellular vesicles of 40-150 nm in size, are secreted by nearly all types of cells and play a dynamic role in intercellular and interorgan communications. These vesicles secreted by source cells contain a variety of biologically active materials such as microRNAs (miRNAs) or proteins, thereby utilizing these cargoes in modifying molecular functionalities of the target cells in the remote tissues. Consequently, several key functions of microenvironmental niches in the tissues are regulated in an exosome-dependent manner. The precise mechanisms by which the exosomes bind and home to different organs remained largely unknown. In recent years, integrins, a large family of cell adhesion molecules, have been revealed to play a crucial role in guiding homing of exosomes to target tissues, as integrins regulate tissue-specific homing of cells. In this regard, it is imperative to experimentally determine the roles played by integrins on the exosomes in their tissue-specific homing. This chapter presents a protocol to investigate exosomal homing regulated by integrins in in vitro and in vivo settings. We focus on ß7 integrin, as its role in mediating the gut-specific homing of lymphocytes has been well established.

Overexpression of S-Adenosylmethionine Synthetase in Recombinant Chlamydomonas for Enhanced Lipid Production.

Microalgae are attracting much attention as promising, eco-friendly producers of bioenergy due to their fast growth, absorption of carbon dioxide from the atmosphere, and production capacity in wastewater and salt water. However, microalgae can only accumulate large quantities of lipid in abiotic stress, which reduces productivity by decreasing cell growth. In this study, the strategy was investigated to increase cell viability and lipid production by overexpressing S-adenosylmethionine (SAM) synthetase (SAMS) in the microalga Chlamydomonas reinhardtii. SAM is a substance that plays an important role in various intracellular biochemical reactions, such as cell proliferation and stress response, and the overexpression of SAMS could allow cells to withstand the abiotic stress and increase productivity. Compared to wild-type C. reinhardtii, recombinant cells overexpressing SAMS grew 1.56-fold faster and produced 1.51-fold more lipids in a nitrogen-depleted medium. Furthermore, under saline-stress conditions, the survival rate and lipid accumulation were 1.56 and 2.04 times higher in the SAMS-overexpressing strain, respectively. These results suggest that the overexpression of SAMS in recombinant C. reinhardtii has high potential in the industrial-scale production of biofuels and various other high-value-added materials.