Endothelial RUNX3 controls LSEC dysfunction and angiocrine LRG1 signaling to prevent liver fibrosis.

BACKGROUND AND AIMS: Liver fibrosis represents a global health burden, given the paucity of approved antifibrotic therapies. Liver sinusoidal endothelial cells (LSECs) play a major gatekeeping role in hepatic homeostasis and liver disease pathophysiology. In early tumorigenesis, runt-related transcription factor 3 (RUNX3) functions as a sentinel; however, its function in liver fibrosis in LSECs remains unclear. This study aimed to investigate the role of RUNX3 as an important regulator of the gatekeeping functions of LSECs and explore novel angiocrine regulators of liver fibrosis. APPROACH AND RESULTS: Mice with endothelial Runx3 deficiency develop gradual and spontaneous liver fibrosis secondary to LSEC dysfunction, thereby more prone to liver injury. Mechanistic studies in human immortalized LSECs and mouse primary LSECs revealed that IL-6/JAK/STAT-3 pathway activation was associated with LSEC dysfunction in the absence of RUNX3. Single-cell RNA sequencing and quantitative RT-PCR revealed that leucine-rich alpha-2-glycoprotein 1 (LRG1) was highly expressed in RUNX3-deficient and dysfunctional LSECs. In in vitro and coculture experiments, RUNX3-depleted LSECs secreted LRG1, which activated hepatic stellate cells via TGFBR1-SMAD2/3 signaling in a paracrine manner. Furthermore, circulating LRG1 levels were elevated in mouse models of liver fibrosis and in patients with fatty liver and cirrhosis. CONCLUSIONS: RUNX3 deficiency in the endothelium induces LSEC dysfunction, LRG1 secretion, and liver fibrosis progression. Therefore, endothelial RUNX3 is a crucial gatekeeping factor in LSECs, and profibrotic angiocrine LRG1 may be a novel target for combating liver fibrosis.

Red2Flpe-SCON: a versatile, multicolor strategy for generating mosaic conditional knockout mice.

Image-based lineage tracing enables tissue turnover kinetics and lineage potentials of different adult cell populations to be investigated. Previously, we reported a genetic mouse model system, Red2Onco, which ectopically expressed mutated oncogenes together with red fluorescent proteins (RFP). This system enabled the expansion kinetics and neighboring effects of oncogenic clones to be dissected. We now report Red2Flpe-SCON: a mosaic knockout system that uses multicolor reporters to label both mutant and wild-type cells. We develop the Red2Flpe mouse line for red clone-specific Flpe expression, as well as the FRT-based SCON (Short Conditional IntrON) method to facilitate tunable conditional mosaic knockouts in mice. We use the Red2Flpe-SCON method to study Sox2 mutant clonal analysis in the esophageal epithelium of adult mice which reveal that the stem cell gene, Sox2, is less essential for adult stem cell maintenance itself, but rather for stem cell proliferation and differentiation.

A dietary commensal microbe enhances antitumor immunity by activating tumor macrophages to sequester iron.

Innate immune cells generate a multifaceted antitumor immune response, including the conservation of essential nutrients such as iron. These cells can be modulated by commensal bacteria; however, identifying and understanding how this occurs is a challenge. Here we show that the food commensal Lactiplantibacillus plantarum IMB19 augments antitumor immunity in syngeneic and xenograft mouse tumor models. Its capsular heteropolysaccharide is the major effector molecule, functioning as a ligand for TLR2. In a two-pronged manner, it skews tumor-associated macrophages to a classically active phenotype, leading to generation of a sustained CD8+ T cell response, and triggers macrophage 'nutritional immunity' to deploy the high-affinity iron transporter lipocalin-2 for capturing and sequestering iron in the tumor microenvironment. This process induces a cycle of tumor cell death, epitope expansion and subsequent tumor clearance. Together these data indicate that food commensals might be identified and developed into 'oncobiotics' for a multi-layered approach to cancer therapy.

Improved predictability of pancreatic ductal adenocarcinoma diagnosis using a blood immune cell biomarker panel developed from bulk mRNA sequencing and single-cell RNA-sequencing.

BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) remains a devastating cancer due to its poor survival rate, early detection, and resectability. This study aimed to determine the peripheral blood mononuclear cell (PBMC) immune biomarkers in patients with PDAC and investigate the PDAC-specific peripheral blood biomarker panel and validate its clinical performance. METHODS: In this prospective, blinded, case-control study, a biomarker panel formula was generated using a development cohort-including healthy controls, patients at high risk of PDAC, and patients with benign pancreatic disease, PDAC, or other gastrointestinal malignancies-and its diagnostic performance was verified using a validation cohort, including patients with ≥ 1 lesion suspected as PDAC on computed tomography (CT). RESULTS: RNA-sequencing of PBMCs from patients with PDAC identified three novel immune cell markers, IL-7R, PLD4, and ID3, as specific markers for PDAC. Regarding the diagnostic performance of the regression formula for the three biomarker panels, the sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were 84.0%, 78.8%, 47.2%, 95.6%, and 79.8%, respectively. Based on the formula scores for the biomarker panel, the false-negative rate (FNR) of the biomarkers was 8% (95% confidence interval [CI] 3.0-13.0), which was significantly lower than that based on CT in the validation cohort (29.2%, 95% CI 20.8-37.6). CONCLUSIONS: The regression formula constructed using three PBMC biomarkers is an inexpensive, rapid, and convenient method that shows clinically useful performance for the diagnosis of PDAC. It aids diagnoses and differential diagnoses of PDAC from pancreatic disease by lowering the FNR compared to CT. Clinical trial registration Clinical Research Information Service, KCT0004614 (08 January 2020).

A synergistic partnership between IL-33/ST2 and Wnt pathway through Bcl-xL drives gastric cancer stemness and metastasis.

ST2 functions as a receptor for the cytokine IL-33. It has been implicated in carcinogenesis. In this study, we sought to mechanistically determine how ST2 and IL-33 function to support cancer stem cell (CSC) activity and drive gastric cancer (GC) pathogenesis. ST2+ subpopulation spontaneously arose during gastric tumorigenesis. A thorough evaluation of ST2 and IL-33 expression in gastric tumors revealed that they show an overlapping expression pattern, notably in poor differentiated GC and metastasis foci. Moreover, their expression levels are clinically correlated to cancer progression. Using a genetic model of CSC-driven gastric carcinogenesis, ST2+ subpopulation displays increased tumorigenicity, chemoresistance and metastatic potentials through increased survival fitness endowed by an elevated MAPK-regulated Bcl-xL. The IL-33/ST2 axis enhances the self-renewal and survival of GC stem cells and organoids. Importantly, we observed a synergistic cooperation between IL-33/ST2 and the canonical Wnt pathway in transactivating Wnt-dependent transcription and supporting CSC activity, a partnership that was abrogated by inhibiting Bcl-xL. Concordant with this, ST2+ subpopulation was targeted by MEK1/2 and Bcl-xL-specific inhibitors. These findings establish ST2 as a functional CSC marker that fortifies the Wnt signal while availing a novel therapeutic strategy to suppress GC progression by targeting the IL-33/ST2/Bcl-xL signaling axis.

Prediction models for early diagnosis of actinomycotic osteomyelitis of the jaw using machine learning techniques: a preliminary study.

BACKGROUND: This study aimed to develop and validate five machine learning models designed to predict actinomycotic osteomyelitis of the jaw. Furthermore, this study determined the relative importance of the predictive variables for actinomycotic osteomyelitis of the jaw, which are crucial for clinical decision-making. METHODS: A total of 222 patients with osteomyelitis of the jaw were analyzed, and Actinomyces were identified in 70 cases (31.5%). Logistic regression, random forest, support vector machine, artificial neural network, and extreme gradient boosting machine learning methods were used to train the models. The models were subsequently validated using testing datasets. These models were compared with each other and also with single predictors, such as age, using area under the receiver operating characteristic (ROC) curve (AUC). RESULTS: The AUC of the machine learning models ranged from 0.81 to 0.88. The performance of the machine learning models, such as random forest, support vector machine and extreme gradient boosting was significantly superior to that of single predictors. Presumed causes, antiresorptive agents, age, malignancy, hypertension, and rheumatoid arthritis were the six features that were identified as relevant predictors. CONCLUSIONS: This prediction model would improve the overall patient care by enhancing prognosis counseling and informing treatment decisions for high-risk groups of actinomycotic osteomyelitis of the jaw.

Transient cAMP elevation during systems consolidation enhances remote contextual fear memory.

Memory is stored in our brains over a temporally graded transition. With time, recently formed memories are transformed into remote memories for permanent storage; multiple brain regions, such as the hippocampus and neocortex, participate in this process. In this study, we aimed to understand the molecular mechanism of systems consolidation of memory and to investigate the brain regions that contribute to this regulation. We first carried out a contextual fear memory test using a transgenic mouse line, which expressed exogenously-derived Aplysia octopamine receptors in the forebrain region, such that, in response to octopamine treatment, cyclic adenosine monophosphate (cAMP) levels could be transiently elevated. From this experiment, we revealed that transient elevation of cAMP levels in the forebrain during systems consolidation led to an enhancement in remote fear memory and increased miniature excitatory synaptic currents in layer II/III of the anterior cingulate cortex (ACC). Furthermore, using an adeno-associated-virus-driven DREADD system, we investigated the specific regions in the forebrain that contribute to the regulation of memory transfer into long-term associations. Our results implied that transient elevation of cAMP levels was induced chemogenetically in the ACC, but not in the hippocampus, and showed a significant enhancement of remote memory. This finding suggests that neuronal activation during systems consolidation through the elevation of cAMP levels in the ACC contributes to remote memory enhancement.

Toll-like receptor mediated inflammation requires FASN-dependent MYD88 palmitoylation.

Toll-like receptor (TLR)/myeloid differentiation primary response protein (MYD88) signaling aggravates sepsis by impairing neutrophil migration to infection sites. However, the role of intracellular fatty acids in TLR/MYD88 signaling is unclear. Here, inhibition of fatty acid synthase by C75 improved neutrophil chemotaxis and increased the survival of mice with sepsis in cecal ligation puncture and lipopolysaccharide-induced septic shock models. C75 specifically blocked TLR/MYD88 signaling in neutrophils. Treatment with GSK2194069 that targets a different domain of fatty acid synthase, did not block TLR signaling or MYD88 palmitoylation. De novo fatty acid synthesis and CD36-mediated exogenous fatty acid incorporation contributed to MYD88 palmitoylation. The binding of IRAK4 to the MYD88 intermediate domain and downstream signal activation required MYD88 palmitoylation at cysteine 113. MYD88 was palmitoylated by ZDHHC6, and ZDHHC6 knockdown decreased MYD88 palmitoylation and TLR/MYD88 activation upon lipopolysaccharide stimulus. Thus, intracellular saturated fatty acid-dependent palmitoylation of MYD88 by ZDHHC6 is a therapeutic target of sepsis.

A Retrospective Study of Patient Outcomes After Temporomandibular Joint Replacement With Alloplastic Total Joint Prosthesis at Massachusetts General Hospital.

PURPOSE: Disorders of the temporomandibular joint (TMJ) occur frequently, with a prevalence of 15 to 18%. Total joint replacement (TJR) surgery is indicated for severe joint damage associated with impaired function, pain, or occlusal change for which other treatments have been unsuccessful. The aim of this study was to assess changes in pain and range of motion (ROM), as well as postoperative complications and comorbidities, in patients receiving TJR surgery at Massachusetts General Hospital (MGH). PATIENTS AND METHODS: This study is a retrospective review that describes the clinical variables in patients after alloplastic TMJ reconstruction performed at MGH from 2000 to 2015. Clinical variables included primary diagnosis; number of previous surgical procedures; comorbidities; preoperative and postoperative pain; preoperative, intraoperative, and postoperative ROM; and complications. RESULTS: Data were obtained from 95 patients undergoing a total of 108 surgical procedures, with an average follow-up period of 4.48 ± 3.38 years. The most common primary indications for TJR were ankylosis (44%) and inflammatory disease (23%). The maximum interincisal opening improved by a mean of 7.7 ± 10.27 mm, and pain decreased by a mean of 1.5 ± 3.29 points on a visual analog scale. Transient facial nerve palsy (25%) was the most common postoperative complication; however, long-term complications were rare. The most frequent comorbidities were psychiatric disorders (56%) and gastrointestinal disease (46%). Psychiatric patients had similar preoperative pain scores (6.0 ± 2.90) but significantly higher postoperative pain scores (4.7 ± 2.58) compared with nonpsychiatric patients. Twenty-eight percent of patients had prior failed TMJ implant materials, specifically Proplast-Teflon (Vitek, Houston, TX). These patients were significantly older (50.4 ± 8.26 years) and had smaller preoperative ROM (21.7 ± 8.85 mm) and smaller postoperative ROM (28.3 ± 9.59 mm). CONCLUSIONS: Patients showed a statistically significant (P < .01) increase in ROM and reduction in pain. TJR is an effective treatment option in patients with limited mouth opening or severe pain.

CD133-induced TM4SF5 expression promotes sphere growth via recruitment and blocking of protein tyrosine phosphatase receptor type F (PTPRF).

CD133 is a surface marker of liver cancer stem cells. Transmembrane 4 L six family member 5 (TM4SF5) promotes sphere growth and circulation. However, it is unknown how CD133 and TM4SF5 cross-talk with each other for cancer stem cell properties. Here, we investigated the significance of inter-relationships between CD133, TM4SF5, CD44, and protein tyrosine phosphatase receptor type F (PTPRF) in a three-dimensional (3D) sphere growth system. We found that CD133 upregulated TM4SF5 and CD44, whereas TM4SF5 and CD44 did not affect CD133 expression. Signaling activity following CD133 phosphorylation caused TM4SF5 expression and sphere growth. TM4SF5 bound to CD133 and promoted c-Src activity for CD133 phosphorylation as a positive feedback loop, leading to CD133-mediated sphere growth that was inhibited by TM4SF5 inhibition or suppression. TM4SF5 also bound PTPRF and promoted paxillin phosphorylation. Decreased sphere growth upon CD133 suppression was recovered by TM4SF5 expression and partially by PTPRF suppression. TM4SF5 inhibition enhanced PTPRF levels and abolished PTPRF suppression-mediated sphere growth. Altogether, CD133-induced TM4SF5 expression and function were important for liver cancer sphere growth and may be a promising target to block metastasis.

Combined or Staged Temporomandibular Joint and Orthognathic Surgery for Patients with Internal Derangement and Dentofacial Deformities.

Patients with internal derangement of the temporomandibular joint and dentofacial deformities need appropriate evaluation for both conditions. Correct diagnosis of internal derangement is vital in determining the correct orthognathic surgery plan, and it is particularly important to differentiate between myofascial dysfunction and intra-articular joint problems. Depending on the stage of internal derangement, patients may need treatment for temporomandibular dysfunction symptomatically, staged, or concurrently with orthognathic surgery.

Evaluation of Andrews' Analysis as a Predictor of Ideal Sagittal Maxillary Positioning in Orthognathic Surgery.

PURPOSE: There is no universally accepted method for determining the ideal sagittal position of the maxilla in orthognathic surgery. In "Element II" of "The Six Elements of Orofacial Harmony," Andrews used the forehead to define the goal maxillary position. The purpose of this study was to compare how well this analysis correlated with postoperative findings in patients who underwent bimaxillary orthognathic surgery planned using other guidelines. The authors hypothesized that the Andrews analysis would more consistently reflect clinical outcomes than standard angular and linear measurements. MATERIALS AND METHODS: This is a retrospective cohort study of patients who had bimaxillary orthognathic surgery and achieved an acceptable esthetic outcome. Patients with no maxillary sagittal movement, obstructive sleep apnea, cleft or craniofacial diagnoses, or who were non-Caucasian were excluded. Treatment plans were developed using photographs, radiographs, and standard cephalometric measurements. The Andrews analysis, measuring the distance from the maxillary incisor to the goal anterior limit line, and standard measurements were applied to end-treatment records. The Andrews analysis was statistically compared with standard methods. RESULTS: There were 493 patients who had orthognathic surgery from 2007 through 2014, and 60 (62% women; mean age, 22.1 ± 6.8 yr) met the criteria for inclusion in this study. The mean Andrews distances were -4.8 ± 2.9 mm for women and -8.6 ± 4.6 mm for men preoperatively and -0.6 ± 2.1 mm for women and -1.9 ± 3.4 mm for men postoperatively. For women, the Andrews analysis was closer to the goal value (0 mm) postoperatively than any standard measurement (P < .001). For men, the linear distance from the A point to a vertical line tangent to the nasion from the McNamara analysis performed best (P < .001), followed by the Andrews analysis. CONCLUSION: The Andrews analysis correlated well with the final esthetic sagittal maxillary position in the present sample, particularly for women, and could be a useful tool for orthognathic surgical planning.

TM4SF5 promotes metastatic behavior of cells in 3D extracellular matrix gels by reducing dependency on environmental cues.

Transmembrane 4 L six family member 5 (TM4SF5) is highly expressed in hepatocellular carcinoma tissues and enhances migration in two-dimensional environments. Here, we investigated how TM4SF5 is involved in diverse pro-metastatic phenotypes in in vivo-like three-dimensional (3D) extracellular matrix gels. TM4SF5-positive cells aggressively formed invasive foci in 3D Matrigel, depending on TM4SF5-mediated signaling activity, cytoskeletal organization, and matrix metallopeptidase (MMP) 2-mediated extracellular remodeling, whereas TM4SF5-null cells did not. The TM4SF5-null cells did, however, form invasive foci in 3D Matrigel following inhibition of Rho-associated protein kinase or addition of collagen I, suggesting that collagen I compensated for TM4SF5 expression. Similarly, TM4SF5-positive cells expressing vascular endothelial-cadherin formed network-like vasculogenic mimicry in 3D Matrigel and collagen I mixture gels, whereas TM4SF5-negative cells in the mixture gels displayed the network structures only upon further treatment with epidermal growth factor. The foci formation also required MMP2-mediated remodeling of the extracellular matrix. Co-cultures exhibited TM4SF5-positive or cancer-associated fibroblasts at the outward edges of TM4SF5-null cell clusters. Compared with TM4SF5-null cells, TM4SF5-positive cells in 3D collagen gels showed a more invasive outgrowth with dramatic invadopodia. These observations suggest that TM4SF5 plays roles in the promotion of diverse metastatic properties with fewer environmental requirements than TM4SF5-negative cells.

Enhanced Systemic Anti-Angiogenic siVEGF Delivery Using PEGylated Oligo-d-arginine.

Angiogenesis mainly mediated by upregulation of vascular endothelial growth factor (VEGF) provides a hallmark of rapidly proliferating tumor cells and an essential component of the tumor growth and microenvironment, making it a targetable process for antitumor therapy. RNA interference (RNAi) provides a very effective tool for developing antitumor therapies; however, its application to date has been hampered due to the lack of efficient small interfering RNA (siRNA) delivery systems in vivo. Here, we report a polymeric gene carrier system based on PEGylation of a cationic cysteine-ended 9-mer arginine oligopeptide (CR9C), which provides effective siRNA systemic delivery and specifically suppresses VEGF (siVEGF). The PEG500-CR9C/siVEGF oligopeptoplex provided improved blood circulation, enhanced protection from serum proteases, reduced uptake in the liver and kidneys, enhanced tumor targeting, and down-regulated intratumoral VEGF level, which comprehensively resulted in improved antitumor efficacy without significant toxicity in vivo. PEG500-CR9C has a great potential for safe and efficient siRNA delivery with diverse applications.

PKCα-mediated phosphorylation of LSD1 is required for presynaptic plasticity and hippocampal learning and memory.

Lysine-specific demethylase 1 (LSD1) is a histone demethylase that participates in transcriptional repression or activation. Recent studies reported that LSD1 is involved in learning and memory. Although LSD1 phosphorylation by PKCα was implicated in circadian rhythmicity, the importance of LSD1 phosphorylation in learning and memory is unknown. In this study, we examined the roles of LSD1 in synaptic plasticity and memory using Lsd1 SA/SA knock-in (KI) mice, in which a PKCα phosphorylation site is mutated. Interestingly, short-term and long-term contextual fear memory as well as spatial memory were impaired in Lsd1 KI mice. In addition, short-term synaptic plasticity, such as paired pulse ratio and post-tetanic potentiation was impaired, whereas long-term synaptic plasticity, including long-term potentiation and long-term depression, was normal. Moreover, the frequency of miniature excitatory postsynaptic current was significantly increased, suggesting presynaptic dysfunction in Lsd1 KI mice. Consistent with this, RNA-seq analysis using the hippocampus of Lsd1 KI mice showed significant alterations in the expressions of presynaptic function-related genes. Intriguingly, LSD1n-SA mutant showed diminished binding to histone deacetylase 1 (HDAC1) compared to LSD1n-WT in SH-SY5Y cells. These results suggest that LSD1 is involved in the regulation of presynaptic gene expression and subsequently regulates the hippocampus-dependent memory in phosphorylation-dependent manner.

Cross-sectional association between testosterone, sex hormone-binding globulin and metabolic syndrome: The Healthy Twin Study.

OBJECTIVES: This study evaluated an association between testosterone, sex hormone-binding globulin (SHBG) and metabolic syndrome (MetS).We also evaluated the genetic and environmental influences on the association. DESIGN: Cross-sectional. SETTING: Community-based study. PARTICIPANTS: A total of 1098 Korean adult men including 139 monozygotic twin pairs. MAIN OUTCOME MEASURE: MetS was defined using the National Cholesterol Education Program-Third Adult Treatment Panel (NCEP ATP III) and International Diabetes Federation (IDF) criteria. The associations between MetS and sex hormones were evaluated using linear mixed model and generalized estimating equation model. RESULTS: After considering covariates such as smoking, alcohol consumption and physical exercises as well as SHBG or testosterone, the risk of MetS defined by NCEP ATP III criteria decreased by 31%, 29%, and 48%, respectively, with 1-standard deviation increase in total testosterone (TT), free testosterone (cFT) and SHBG. Similar findings were revealed with IDF criteria. Metabolic component specific analysis showed that sex hormones were inversely associated with several components of MetS: TT with abdominal obesity, low high-density lipoprotein cholesterol (HDL-C) and high blood pressure; cFT with abdominal obesity and high blood pressure; SHBG with all components except high blood pressure. Cotwin control analysis found an inverse correlation between within-pair differences in testosterone and SHBG levels and within-pair differences in waist circumference only. CONCLUSION: Both testosterone and SHBG were inversely associated with MetS although the inverse associations with the sex hormones were not consistently found across individual metabolic components. Findings from cotwin analysis suggest a significant contribution of unshared unique environmental effect to the association between testosterone and SHBG and abdominal obesity.

Dynamic and coordinated single-molecular interactions at TM4SF5-enriched microdomains guide invasive behaviors in 2- and 3-dimensional environments.

Membrane proteins sense extracellular cues and transduce intracellular signaling to coordinate directionality and speed during cellular migration. They are often localized to specific regions, as with lipid rafts or tetraspanin-enriched microdomains; however, the dynamic interactions of tetraspanins with diverse receptors within tetraspanin-enriched microdomains on cellular surfaces remain largely unexplored. Here, we investigated effects of tetraspan(in) TM4SF5 (transmembrane 4 L6 family member 5)-enriched microdomains (T5ERMs) on the directionality of cell migration. Physical association of TM4SF5 with epidermal growth factor receptor (EGFR) and integrin α5 was visualized by live fluorescence cross-correlation spectroscopy and higher-resolution microscopy at the leading edge of migratory cells, presumably forming TM4SF5-enriched microdomains. Whereas TM4SF5 and EGFR colocalized at the migrating leading region more than at the rear, TM4SF5 and integrin α5 colocalized evenly throughout cells. Cholesterol depletion and disruption in TM4SF5 post-translational modifications, including N-glycosylation and palmitoylation, altered TM4SF5 interactions and cellular localization, which led to less cellular migration speed and directionality in 2- or 3-dimensional conditions. TM4SF5 controlled directional cell migration and invasion, and importantly, these TM4SF5 functions were dependent on cholesterol, TM4SF5 post-translational modifications, and EGFR and integrin α5 activity. Altogether, we showed that TM4SF5 dynamically interacted with EGFR and integrin α5 in migratory cells to control directionality and invasion.-Kim, H.-J., Kwon, S., Nam, S. H., Jung, J. W., Kang, M., Ryu, J., Kim, J. E., Cheong, J.-G., Cho, C. Y., Kim, S., Song, D.-G., Kim, Y.-N., Kim, T. Y., Jung, M.-K., Lee, K.-M., Pack, C.-G., Lee, J. W. Dynamic and coordinated single-molecular interactions at TM4SF5-enriched microdomains guide invasive behaviors in 2- and 3-dimensional environments.

Differential regulation of cellular functions by the C-termini of transmembrane 4 L six family proteins in 2- or 3-dimensional environment.

The transmembrane 4 L six family proteins TM4SF1, TM4SF4, and TM4SF5 share 40-50% overall sequence identity, but their C-terminus identity is limited. It may be likely that the C-termini of the members are important and unique for own regulatory functions. We thus examined how the TM4SF5 C-terminus affected cellular functions differentially from other family members. Using colon cancer cells expressing wildtype (WT), C-terminus-deleted, or chimeric mutants, diverse cellular functions were explored in 2-dimensional (2D) and 3-dimensional (3D) condition. The C-termini of the proteins were relatively comparable with respect to 2D cell proliferation, although each C-terminal-deletion mutant exhibited increased proliferation relative to the WT. Using chimeric constructs, we found that the TM4SF5 C-terminus was critical for regulating the diverse metastatic functions of TM4SF5, and could positively replace the C-termini of other family members. Replacement of the TM4SF1 or TM4SF4 C-terminus with that of TM4SF5 increased spheroids growth, transwell migration, and invasive dissemination from spheroids in 3D collagen gels. TM4SF5-mediated effects required its extracellular loop 2 linked to the C-terminus via the transmembrane domain 4, with causing c-Src activation. Altogether, the C-terminus of TM4SF5 appears to mediate pro-migratory roles, depending on a structural relay from the second extracellular loop to the C-terminus.

Cytokine-like Activity of Liver Type Fatty Acid Binding Protein (L-FABP) Inducing Inflammatory Cytokine Interleukin-6.

It has been reported that fatty acid binding proteins (FABPs) do not act only as intracellular mediators of lipid responses but also have extracellular functions. This study aimed to investigate whether extracellular liver type (L)-FABP has a biological activity and to determined serum L-FABP levels in patients with end-stage renal disease (ESRD). We isolated L-FABP complementary deoxyribonucleic acid (cDNA) from the Huh7 human hepatocarcinoma cell line and expressed the recombinant L-FABP protein in Escherichia coli. A549 lung carcinoma and THP-1 monocytic cells were stimulated with the human recombinant L-FABP. Human whole blood cells were also treated with the human recombinant L-FABP or interleukin (IL)-1α. IL-6 levels were measured in cell culture supernatants using IL-6 enzyme-linked immunosorbent assay (ELISA). Human recombinant L-FABP induced IL-6 in a dose-dependent manner in A549, THP-1 cells, and whole blood cells. The blood samples of healthy volunteers and patients with ESRD were taken after an overnight fast. The serum levels of L-FABP in healthy volunteers and ESRD patients were quantified with L-FABP ELISA. The values of L-FABP in patients with ESRD were significantly lower than those in the control group. Our results demonstrated the biological activity of L-FABP in human cells suggesting L-FABP can be a mediator of inflammation.

Impaired learning and memory in CD38 null mutant mice.

CD38 is an enzyme that catalyzes the formation of cyclic ADP ribose and nicotinic acid adenine dinucleotide phosphate, both of which are involved in the mobilization of Ca(2+) from intracellular stores. Recently, CD38 has been shown to regulate oxytocin release from hypothalamic neurons. Importantly, CD38 mutations are associated with autism spectrum disorders (ASD) and CD38 knockout (CD38(-/-)) mice display ASD-like behavioral phenotypes including deficient parental behavior and poor social recognition memory. Although ASD and learning deficits commonly co-occur, the role of CD38 in learning and memory has not been investigated. We report that CD38(-/-) mice show deficits in various learning and memory tasks such as the Morris water maze, contextual fear conditioning, and the object recognition test. However, either long-term potentiation or long-term depression is not impaired in the hippocampus of CD38(-/-) mice. Our results provide convincing evidence that CD38(-/-) mice show deficits in various learning and memory tasks including spatial and non-spatial memory tasks. Our data demonstrate that CD38 is critical for regulating hippocampus-dependent learning and memory without modulating synaptic plasticity.