Staging Laparoscopy is Underutilized in the Management of Gastric Adenocarcinoma.

BACKGROUND: Staging laparoscopy (SL) with peritoneal lavage is usually performed on a separate day from the planned resection and is recommended in patients with gastric adenocarcinoma as it can identify radiographically occult metastases and malignant cytology, thus altering prognosis and treatment. SL can be done on the same day as planned resection (SLSR) or with delayed resection (SLDR). The purpose of this study was to determine utilization of SL and factors associated with SLSR and SLDR, among patients diagnosed with gastric adenocarcinoma. METHODS: SEER-Medicare linked data were used to identify patients diagnosed with gastric adenocarcinoma from 2004 through 2013. SL were defined as a laparoscopy that occurred up to 3 months postdiagnosis. Multivariate logistic regression was used to identify factors associated with the utilization of SLSR and SLDR. RESULTS: Of the 5610 patients with gastric adenocarcinoma who underwent a surgical procedure, 733 (13%) had a SL. Utilization of SL increased annually from 6.4% to 22.2% (p < 0.01). Receipt of SL was associated with patient demographics, tumor location, and treatment at a National Cancer Institute (NCI) Designated Cancer Center (CC). Of the 733 patients who underwent SL, 475 (65%) received further surgical procedures; 367 (77%) underwent SLSR, while 108 patients (23%) underwent SLDR. Compared with SLSR, SLDR was more common among patients who were younger, treated at an NCI-Designated CC and had proximal tumors. CONCLUSIONS: SL for optimal preoperative staging remains underutilized in the management of gastric adenocarcinoma. Expanded use of laparoscopy as a distinct procedure could minimize unnecessary interventions.

Outcomes for Correction of Long-Gap Esophageal Atresia: A 22-Year Experience.

BACKGROUND: Long-gap esophageal atresia (LGEA) precludes immediate primary repair. When delayed primary esophagoesophagostomy (DPE) is not feasible, a reverse gastric tube (RGT) is a potential salvage option. The purpose of this study was to determine if DPE and RGT had both similar short-term and long-term outcomes. METHODS: A retrospective review of all EA patients from 1994 to 2016 was undertaken. Data were stratified by surgical management (DPE versus RGT). Baseline demographics, operative information, postoperative management, and complications were analyzed. Descriptive statistics were used and P-values <0.05 were considered statistically significant. RESULTS: Two hundred and eighteen patients with EA were treated during this period; 37/218 (17%) had LGEA. Mean gap length was 3.3 ± 1.2 cm. Thirty-three patients underwent some form of repair, all of which were managed initially with a gastrostomy tube feeds. Twenty-five patients underwent DPE with 89% of these never requiring revision, and 86% having excellent function with long-term follow-up. In eight patients, esophageal length was never adequate for DPE; therefore, six were reconstructed with RGT, and two underwent gastric transposition. There were no significant differences in complications, revisions, ventilator days, overall length of stay, weight percentiles, or conduit function between children undergoing RGT compared with DPE at a mean follow-up of 5.5 years. CONCLUSIONS: Surgical treatment of LGEA is complex, and controversy exists regarding the optimal repair method when DPE is not feasible. In this series, DPE after gastrostomy tube feeds often allowed for sufficient esophageal lengthening with satisfactory long-term esophageal function. However, when adequate length for DPE was not attainable, these data suggest that RGT is a viable conduit with favorable postoperative outcomes.

T-Cell Transfer Therapy Targeting Mutant KRAS in Cancer.

We identified a polyclonal CD8+ T-cell response against mutant KRAS G12D in tumor-infiltrating lymphocytes obtained from a patient with metastatic colorectal cancer. We observed objective regression of all seven lung metastases after the infusion of approximately 1.11×1011 HLA-C*08:02-restricted tumor-infiltrating lymphocytes that were composed of four different T-cell clonotypes that specifically targeted KRAS G12D. However, one of these lesions had progressed on evaluation 9 months after therapy. The lesion was resected and found to have lost the chromosome 6 haplotype encoding the HLA-C*08:02 class I major histocompatibility complex (MHC) molecule. The loss of expression of this molecule provided a direct mechanism of tumor immune evasion. Thus, the infusion of CD8+ cells targeting mutant KRAS mediated effective antitumor immunotherapy against a cancer that expressed mutant KRAS G12D and HLA-C*08:02.

Physical Examination is the Best Predictor of the Need for Abdominal Surgery in Children Following Motor Vehicle Collision.

BACKGROUND: Exploratory laparotomy in children after motor vehicle collision (MVC) is rare. In the absence of definitive hemorrhage or free abdominal air on radiographic imaging, predictors for operative exploration are conflicting. OBJECTIVE: The purpose of this study was to explore objective findings that may aid in determining which children require operative abdominal exploration after MVC. METHODS: Data from 2010-2014 at an American College of Surgeons-certified level 1 pediatric trauma center were retrospectively reviewed. Demographics, vital signs, laboratory data, radiologic studies, operative records, associated injuries, and outcomes were analyzed and p < 0.05 was considered statistically significant. RESULTS: Eight hundred sixty-two patients 0-18 years of age presented to the hospital after an MVC during the study period. Seventeen patients (2.0%) required abdominal exploration and all were found to have intraabdominal injuries. Respiratory rate was the only vital sign that was significantly altered (p = 0.04) in those who required abdominal surgery compared with those who did not. Physical examination findings, such as the seat belt sign, abdominal bruising, abdominal wound, and abdominal tenderness, were present significantly more frequently in those requiring abdominal surgery (p < 0.0001). Each finding had a negative predictive value for the need for operative exploration of at least 0.98. There were no significant differences in trauma laboratory values or radiographic findings between the 2 groups. CONCLUSION: Data from this study solidify the relationship between specific physical examination findings and the need for abdominal exploration after MVC in children. In addition, these data suggest that a lack of the seat belt sign, abdominal bruising, abdominal wounds, or abdominal tenderness are individually predictive of patients who will not require surgical intervention.

Kidney regeneration: common themes from the embryo to the adult.

The vertebrate kidney has an inherent ability to regenerate following acute damage. Successful regeneration of the injured kidney requires the rapid replacement of damaged tubular epithelial cells and reconstitution of normal tubular function. Identifying the cells that participate in the regeneration process as well as the molecular mechanisms involved may reveal therapeutic targets for the treatment of kidney disease. Renal regeneration is associated with the expression of genetic pathways that are necessary for kidney organogenesis, suggesting that the regenerating tubular epithelium may be "reprogrammed" to a less-differentiated, progenitor state. This review will highlight data from various vertebrate models supporting the hypothesis that nephrogenic genes are reactivated as part of the process of kidney regeneration following acute kidney injury (AKI). Emphasis will be placed on the reactivation of developmental pathways and how our understanding of the resulting regeneration process may be enhanced by lessons learned in the embryonic kidney.

Inhibition of histone deacetylase expands the renal progenitor cell population.

One of the first hallmarks of kidney regeneration is the reactivation of genes normally required during organogenesis. Identification of chemicals with the potential to enhance this reactivation could therapeutically promote kidney regeneration. Here, we found that 4-(phenylthio)butanoic acid (PTBA) expanded the expression domains of molecular markers of kidney organogenesis in zebrafish. PTBA exhibits structural and functional similarity to the histone deacetylase (HDAC) inhibitors 4-phenylbutanoic acid and trichostatin A; treatment with these HDAC inhibitors also expanded the renal progenitor cell population. Analyses in vitro and in vivo confirmed that PTBA functions as an inhibitor of HDAC activity. Furthermore, PTBA-mediated renal progenitor cell expansion required retinoic acid signaling. In summary, these results support a mechanistic link among renal progenitor cells, HDAC, and the retinoid pathway. Whether PTBA holds promise as a therapeutic agent to promote renal regeneration requires further study.

Trends in Major Gastrectomy for Cancer: Frequency and Outcomes.

BACKGROUND: Declining incidence of gastric cancer in the USA has presumably resulted in lower rates of major gastrectomy for cancer. The impact on perioperative outcomes remains undefined. The aims of this study were to characterize national trends in frequency of major gastrectomy for cancer, identify factors associated with in-hospital mortality, and examine outcome disparities by race/ethnicity. METHODS: Nationwide inpatient sample data from 1993 to 2013 were queried for procedural and diagnostic codes (ICD-9) relating to total and partial gastrectomy procedures. Gastric resections for cancer were compared to those for peptic ulcer disease for reference. Patient demographics, comorbidity score, mortality, and hospital characteristics were recorded as covariates. RESULTS: A significant decrease in annual rates of partial and total gastrectomy was observed from 1993 to 2013 (p < 0.0001). The change in absolute number and percent decline was greater for partial gastrectomy (- 39.3%) than total gastrectomy (- 19%). There was a 34.0% decrease in gastrectomy for cancer in Whites and a 61.2% increase among Hispanic patients over two decades. In-hospital mortality also significantly decreased over the study period (7.7% to 2.7%). Factors associated with lower mortality rates included male sex and treatment at urban teaching hospitals. Analysis of trends revealed that gastrectomy for cancer was performed with increasing frequency at urban teaching hospitals. CONCLUSIONS: The frequency of major gastric resections in the USA has declined over two decades. Overall, in-hospital mortality rates also have decreased significantly. Declining in-hospital mortality after gastrectomy for cancer is associated with more frequent treatment at urban teaching hospitals.

Recognition of human gastrointestinal cancer neoantigens by circulating PD-1+ lymphocytes.

Tumor-resident lymphocytes can mount a response against neoantigens expressed in microsatellite-stable gastrointestinal (GI) cancers, and adoptive transfer of neoantigen-specific lymphocytes has demonstrated antitumor activity in selected patients. However, whether peripheral blood could be used as an alternative minimally invasive source to identify lymphocytes targeting neoantigens in patients with GI cancer with relatively low mutation burden is unclear. We used a personalized high-throughput screening strategy to investigate whether PD-1 expression in peripheral blood could be used to identify CD8+ or CD4+ lymphocytes recognizing neoantigens identified by whole-exome sequencing in 7 patients with GI cancer. We found that neoantigen-specific lymphocytes were preferentially enriched in the CD8+PD-1+/hi or CD4+PD-1+/hi subsets, but not in the corresponding bulk or PD-1- fractions. In 6 of 7 individuals analyzed we identified circulating CD8+ and CD4+ lymphocytes targeting 6 and 4 neoantigens, respectively. Moreover, neoantigen-reactive T cells and a T cell receptor (TCR) isolated from the CD8+PD-1+ subsets recognized autologous tumor, albeit at reduced levels, in 2 patients with available cell lines. These data demonstrate the existence of circulating T cells targeting neoantigens in GI cancer patients and provide an approach to generate enriched populations of personalized neoantigen-specific lymphocytes and isolate TCRs that could be exploited therapeutically to treat cancer.

Unique Neoantigens Arise from Somatic Mutations in Patients with Gastrointestinal Cancers.

Immunotherapies can mediate regression of human tumors with high mutation rates, but responses are rarely observed in patients with common epithelial cancers. This raises the question of whether patients with these common cancers harbor T lymphocytes that recognize mutant proteins expressed by autologous tumors that may represent ideal targets for immunotherapy. Using high-throughput immunologic screening of mutant gene products identified via whole-exome sequencing, we identified neoantigen-reactive tumor-infiltrating lymphocytes (TIL) from 62 of 75 (83%) patients with common gastrointestinal cancers. In total, 124 neoantigen-reactive TIL populations were identified, and all but one of the neoantigenic determinants were unique. The results of in vitro T-cell recognition assays demonstrated that 1.6% of the gene products encoded by somatic nonsynonymous mutations were immunogenic. These findings demonstrate that the majority of common epithelial cancers elicit immune recognition and open possibilities for cell-based immunotherapies for patients bearing these cancers. SIGNIFICANCE: TILs cultured from 62 of 75 (83%) patients with gastrointestinal cancers recognized neoantigens encoded by 1.6% of somatic mutations expressed by autologous tumor cells, and 99% of the neoantigenic determinants appeared to be unique and not shared between patients.This article is highlighted in the In This Issue feature, p. 983.

Temporal Variations in Pediatric Trauma: Rationale for Altered Resource Utilization.

Trauma is a major cause of morbidity and mortality in the pediatric population. However, temporal variations of trauma have not been well characterized and may have implications for appropriate allocation of hospital resources. Data from patients evaluated at an ACS-verified Level I pediatric trauma center between 2011 and 2015 were retrospectively analyzed. Date and time of injury, type of injury (blunt vs penetrating), and postemergency department disposition were reviewed. To assess temporal trends, heatmaps were constructed and a mixed poisson regression model was used to assess statistical significance. Pediatric trauma from blunt and penetrating injuries occurred at significantly higher rates between the hours of 1800 and 0100, on weekends compared with weekdays, and from May to August compared with November to February. These data provide useful information for hospital resource utilization. The emergency department, operating room, and intensive care unit should be prepared for increased trauma-related volume between May and August, weekends, and evening hours by appropriately increasing staff volume and resource availability.

Functional analysis of the hepatic HMG-CoA reductase promoter by in vivo electroporation.

HMG-CoA reductase (HMGR) catalyzes the rate-controlling step in cholesterol production. This enzyme is highly expressed in the liver, where it is subject to extensive hormonal and dietary regulation. Although much is known about the regulation of the HMGR promoter in cultured cells, this issue has not been directly addressed in liver. The technique of in vivo electroporation was utilized to perform the first functional analysis of the HMGR promoter in live animals. Analysis of a series of deletion constructs showed that deletion of the region containing the cyclic AMP response element (CRE) at -104 to -96 and an NF-Y site at -70 to -65 resulted in marked reduction of promoter activity. Sterol regulation of this promoter was investigated by raising tissue cholesterol levels by feeding cholesterol and by decreasing them through administration of a statin (lovastatin). Using this approach, we found that HMGR promoter constructs were sterol responsive in live animals, adding in vivo relevance to previous findings in cultured cells. We also conclude that in vivo electroporation is a convenient and powerful technique for the analysis of promoter elements in the livers of live animals.

MAGE-A is More Highly Expressed Than NY-ESO-1 in a Systematic Immunohistochemical Analysis of 3668 Cases.

Two cancer testis antigens, the New York esophageal squamous cell carcinoma-1 (NY-ESO-1) and the melanoma-antigen family A (MAGE-A), represent promising immunotherapy targets due to the low expression of these antigens in nonmalignant tissue. To assess overexpression patterns in various cancers, we performed a systematic immunohistochemical analysis for NY-ESO-1 and MAGE-A on tissue array samples of 3668 common epithelial carcinomas (CA) and germ cell tumors of high prevalence and mortality. Here, we find significantly higher expression of MAGE-A (>50% on tumor cells) compared with NY-ESO-1 in several CAs including cutaneous squamous cell carcinomas (SCC) (52.8%/2.8%), esophageal SCC (50%/0%), head and neck SCC (41.1%/<1%), bladder urothelial CA (40.4%/8.3%), cervical/anal SCC (37.5%/0%), lung SCC (34%/3.8%), lung adenocarcinomas (27.6%/3.9%), ovarian CA (26.4%/3.6%), endometrial CA (26.3%/1.3%), lung small cell CA (24.4%/2.4%), gastric adenocarcinomas (20%/4%), breast mucinous CA (19.3%/0%), hepatocellular CA (18.8%/1.2%), breast infiltrating ductal CA (16.4%/1.8%), colorectal adenocarcinomas (10.7%/<1%), cholangiocarcinomas (9.8%/0%), thymic CA (9%/4.5%), and mesotheliomas (7.9%/<1%). Furthermore, high expression of MAGE-A, but not NY-ESO-1, was seen in whole slide evaluations of an independent cohort of metastatic SCC (45.5%/3.6%) and metastatic CA (13.5%/0%) of various primaries with significantly higher expression of MAGE-A in metastatic SCC compared with other metastatic CA. MAGE-A is also more highly expressed in germ cell tumors, seminomas (69%/3.5%) and nonseminomas (40.1%/4.7%). In summary, MAGE-A is more highly expressed than NY-ESO-1 in a majority of human malignancies, and targeting MAGE-A may benefit a large number of patients.

Expression of MAGE-A and NY-ESO-1 in Primary and Metastatic Cancers.

Melanoma-associated antigen-A (MAGE-A) and New York esophageal squamous cell cancer-1 (NY-ESO-1) are 2 cancer testis antigens (CTA) demonstrating potential for use in targeted immunotherapy. Clinical trials in melanoma and synovial sarcomas targeting these antigens in immune-based therapies have demonstrated durable tumor regression. Although protein expression of NY-ESO-1 has been assessed in a variety of cancer types, the expression of MAGE-A has not been studied in depth. In this study we analyzed MAGE-A and NY-ESO-1 expression in 314 melanoma specimens from 301 melanoma patients, 38 patients with squamous cell cancers and 111 patients with adenocarcinomas. Our results demonstrated higher expression of MAGE-A compared with NY-ESO-1 in melanomas (32% vs. 13%) and squamous cell carcinomas (45% vs. 7.9%), and higher expression of both CTAs in metastatic versus primary tumors. CTA expression in adenocarcinomas was low (MAGE-A: 10%, NY-ESO-1: 0.9%). In addition, we looked at concordance of expression among metastatic melanoma lesions within the same patient and found concordant expression in 38 of 47 patients for MAGE-A and 43 of 47 patients for NY-ESO-1. Our study demonstrated that the MAGE-A family may be of greater utility than NY-ESO-1 for targeted immunotherapy in a variety of cancer histologies, in particular metastatic melanomas and squamous cell carcinomas.

Synergy between loss of NF1 and overexpression of MYCN in neuroblastoma is mediated by the GAP-related domain.

Earlier reports showed that hyperplasia of sympathoadrenal cell precursors during embryogenesis in Nf1-deficient mice is independent of Nf1's role in down-modulating RAS-MAPK signaling. We demonstrate in zebrafish that nf1 loss leads to aberrant activation of RAS signaling in MYCN-induced neuroblastomas that arise in these precursors, and that the GTPase-activating protein (GAP)-related domain (GRD) is sufficient to suppress the acceleration of neuroblastoma in nf1-deficient fish, but not the hypertrophy of sympathoadrenal cells in nf1 mutant embryos. Thus, even though neuroblastoma is a classical "developmental tumor", NF1 relies on a very different mechanism to suppress malignant transformation than it does to modulate normal neural crest cell growth. We also show marked synergy in tumor cell killing between MEK inhibitors (trametinib) and retinoids (isotretinoin) in primary nf1a-/- zebrafish neuroblastomas. Thus, our model system has considerable translational potential for investigating new strategies to improve the treatment of very high-risk neuroblastomas with aberrant RAS-MAPK activation.

Loss of neurofibromin Ras-GAP activity enhances the formation of cardiac blood islands in murine embryos.

Type I neurofibromatosis (NF1) is caused by mutations in the NF1 gene encoding neurofibromin. Neurofibromin exhibits Ras GTPase activating protein (Ras-GAP) activity that is thought to mediate cellular functions relevant to disease phenotypes. Loss of murine Nf1 results in embryonic lethality due to heart defects, while mice with monoallelic loss of function mutations or with tissue-specific inactivation have been used to model NF1. Here, we characterize previously unappreciated phenotypes in Nf1-/- embryos, which are inhibition of hemogenic endothelial specification in the dorsal aorta, enhanced yolk sac hematopoiesis, and exuberant cardiac blood island formation. We show that a missense mutation engineered into the active site of the Ras-GAP domain is sufficient to reproduce ectopic blood island formation, cardiac defects, and overgrowth of neural crest-derived structures seen in Nf1-/-embryos. These findings demonstrate a role for Ras-GAP activity in suppressing the hemogenic potential of the heart and restricting growth of neural crest-derived tissues.

Modulation of cAMP and ras signaling pathways improves distinct behavioral deficits in a zebrafish model of neurofibromatosis type 1.

Neurofibromatosis type 1 (NF1) is a common autosomal-dominant disorder associated with attention deficits and learning disabilities. The primary known function of neurofibromin, encoded by the NF1 gene, is to downregulate Ras activity. We show that nf1-deficient zebrafish exhibit learning and memory deficits and that acute pharmacological inhibition of downstream targets of Ras (MAPK and PI3K) restores memory consolidation and recall but not learning. Conversely, acute pharmacological enhancement of cAMP signaling restores learning but not memory. Our data provide compelling evidence that neurofibromin regulates learning and memory by distinct molecular pathways in vertebrates and that deficits produced by genetic loss of function are reversible. These findings support the investigation of cAMP signaling enhancers as a companion therapy to Ras inhibition in the treatment of cognitive dysfunction in NF1.

Zebrafish neurofibromatosis type 1 genes have redundant functions in tumorigenesis and embryonic development.

Neurofibromatosis type 1 (NF1) is a common, dominantly inherited genetic disorder that results from mutations in the neurofibromin 1 (NF1) gene. Affected individuals demonstrate abnormalities in neural-crest-derived tissues that include hyperpigmented skin lesions and benign peripheral nerve sheath tumors. NF1 patients also have a predisposition to malignancies including juvenile myelomonocytic leukemia (JMML), optic glioma, glioblastoma, schwannoma and malignant peripheral nerve sheath tumors (MPNSTs). In an effort to better define the molecular and cellular determinants of NF1 disease pathogenesis in vivo, we employed targeted mutagenesis strategies to generate zebrafish harboring stable germline mutations in nf1a and nf1b, orthologues of NF1. Animals homozygous for loss-of-function alleles of nf1a or nf1b alone are phenotypically normal and viable. Homozygous loss of both alleles in combination generates larval phenotypes that resemble aspects of the human disease and results in larval lethality between 7 and 10 days post fertilization. nf1-null larvae demonstrate significant central and peripheral nervous system defects. These include aberrant proliferation and differentiation of oligodendrocyte progenitor cells (OPCs), dysmorphic myelin sheaths and hyperplasia of Schwann cells. Loss of nf1 contributes to tumorigenesis as demonstrated by an accelerated onset and increased penetrance of high-grade gliomas and MPNSTs in adult nf1a(+/-); nf1b(-/-); p53(e7/e7) animals. nf1-null larvae also demonstrate significant motor and learning defects. Importantly, we identify and quantitatively analyze a novel melanophore phenotype in nf1-null larvae, providing the first animal model of the pathognomonic pigmentation lesions of NF1. Together, these findings support a role for nf1a and nf1b as potent tumor suppressor genes that also function in the development of both central and peripheral glial cells as well as melanophores in zebrafish.

Cyclin D1 splice variants: polymorphism, risk, and isoform-specific regulation in prostate cancer.

PURPOSE: Alternative CCND1 splicing results in cyclin D1b, which has specialized, protumorigenic functions in prostate not shared by the cyclin D1a (full length) isoform. Here, the frequency, tumor relevance, and mechanisms controlling cyclin D1b were challenged. EXPERIMENTAL DESIGN: First, relative expression of both cyclin D1 isoforms was determined in prostate adenocarcinomas. Second, relevance of the androgen axis was determined. Third, minigenes were created to interrogate the role of the G/A870 polymorphism (within the splice site), and findings were validated in primary tissue. Fourth, the effect of G/A870 on cancer risk was assessed in two large case-control studies. RESULTS: Cyclin D1b is induced in tumors, and a significant subset expressed this isoform in the absence of detectable cyclin D1a. Accordingly, the isoforms showed noncorrelated expression patterns, and hormone status did not alter splicing. Whereas G/A870 was not independently predictive of cancer risk, A870 predisposed for transcript-b production in cells and in normal prostate. The influence of A870 on overall transcript-b levels was relieved in tumors, indicating that aberrations in tumorigenesis likely alter the influence of the polymorphism. CONCLUSIONS: These studies reveal that cyclin D1b is specifically elevated in prostate tumorigenesis. Cyclin D1b expression patterns are distinct from that observed with cyclin D1a. The A870 allele predisposes for transcript-b production in a context-specific manner. Although A870 does not independently predict cancer risk, tumor cells can bypass the influence of the polymorphism. These findings have major implications for the analyses of D-cyclin function in the prostate and provide the foundation for future studies directed at identifying potential modifiers of the G/A870 polymorphism.

High density lipoprotein-associated sphingosine 1-phosphate promotes endothelial barrier function.

High density lipoproteins (HDL) are major plasma carriers of sphingosine 1-phosphate (S1P). Here we show that HDL increases endothelial barrier integrity as measured by electric cell substrate impedance sensing. S1P was implicated as the mediator in this process through findings showing that pertussis toxin, an inhibitor of Gi-coupled S1P receptors, as well as antagonists of the S1P receptor, S1P1, inhibited barrier enhancement by HDL. Additional findings show that HDL stimulates endothelial cell activation of Erk1/2 and Akt, signaling pathway intermediates that have been implicated in S1P-dependent endothelial barrier activity. HDL was also found to promote endothelial cell motility, a process that may also relate to endothelial barrier function in the context of a vascular injury response. The effects of HDL on endothelial cell Erk1/2 and Akt activation and motility were suppressed by pertussis toxin and S1P1 antagonists. However, both HDL-induced barrier enhancement and HDL-induced motility showed a greater dependence on Akt activation as compared with Erk1/2 activation. Together, the findings indicate that HDL has endothelial barrier promoting activities, which are attributable to its S1P component and signaling through the S1P1/Akt pathway.