Omega-3 Polyunsaturated Fatty Acids Suppress the Cystic Lesion Formation of Peritoneal Endometriosis in Transgenic Mouse Models

Omega-3 polyunsaturated fatty acids (omega-3 PUFAs) play a role in controlling pathological inflammatory reactions. Endometriosis is characterized by the presence of endometrial tissue on the peritoneum and an exaggerated inflammatory environment around ectopic tissues. Here peritoneal endometriosis was reproduced using a mouse model in which murine endometrial fragments were inoculated into the peritoneal cavity of mice. Fat-1 mice, in which omega-6 can be converted to omega-3 PUFAs, or wild type mice, in which it cannot, were used for the endometriosis model to address the actions of omega-3 PUFAs on the development of endometriotic lesions. The number and weight of cystic endometriotic lesions in fat-1 mice two weeks after inoculation were significantly less than half to those of controls. Mediator lipidomics revealed that cystic endometriotic lesions and peritoneal fluids were abundant in 12/15-hydroxyeicosapentaenoic acid (12/15-HEPE), derived from eicosapentaenoic acid (EPA), and their amount in fat-1 mice was significantly larger than that in controls. 12/15-Lipoxygenase (12/15-LOX)-knockout (KO) and control mice with or without EPA administration were assessed for the endometriosis model. EPA administration decreased the number of lesions in controls but not in 12/15-LOX-KO mice. The peritoneal fluids in EPA-fed 12/15-LOX-KO mice contained reduced levels of EPA metabolites such as 12/15-HEPE and EPA-derived resolvin E3 even after EPA administration. cDNA microarrays of endometriotic lesions revealed that Interleukin-6 (IL-6) expression in fat-1 mice was significantly lower than that in controls. These results suggest that both endogenous and exogenous EPA-derived PUFAs protect against the development of endometriosis through their anti-inflammatory effects and, in particular, the 12/15-LOX-pathway products of EPA may be key mediators to suppress endometriosis

Increased tissue levels of omega-3 polyunsaturated fatty acids prevents pathological preterm birth

Omega-3 polyunsaturated fatty acids such as eicosapentaenoic acid (EPA) have anti-inflammatory effects. Preterm birth is an important problem in modern obstetrics and one of the main causes is an inflammation. We here showed that abundance of omega-3 fatty acids reduced the incidence of preterm birth induced by LPS with fat-1 mice, capable of converting omega-6 to omega-3 fatty acids. We also indicated that the gene expression of IL-6 and IL-1β in uteruses and the number of cervical infiltrating macrophages were reduced in fat-1 mice. The analyses of lipid metabolomics showed the high level of 18-hydroxyeicosapentaenoate in fat-1 mice, which was derived from EPA and was metabolized to anti-inflammatory product named resolvin E3 (RvE3). We finally showed that the administration of RvE3 to LPS-exposed pregnant wild type mice lowered the incidence of preterm birth. Our data suggest that RvE3 could be a potential new therapeutic for the prevention of preterm birth

CD1d, a Sentinel Molecule Bridging Innate and Adaptive Immunity, Is Downregulated by the Human Papillomavirus (HPV) E5 Protein: a Possible Mechanism for Immune Evasion by HPV▿

CD1d and CD1d-restricted natural killer T (NKT) cells serve as a natural bridge between innate and adaptive immune responses to microbes. CD1d downregulation is utilized by a variety of microbes to evade immune detection. We demonstrate here that CD1d is downregulated in human papillomavirus (HPV)-positive cells in vivo and in vitro. CD1d immunoreactivity was strong in HPV-negative normal cervical epithelium but absent in HPV16-positive CIN1 and HPV6-positive condyloma lesions. We used two cell lines for in vitro assay; one was stably CD1d-transfected cells established from an HPV-negative cervical cancer cell line, C33A (C33A/CD1d), and the other was normal human vaginal keratinocyte bearing endogenous CD1d (Vag). Flow cytometry revealed that cell surface CD1d was downregulated in both C33A/CD1d and Vag cells stably transfected with HPV6 E5 and HPV16 E5. Although the steady-state levels of CD1d protein decreased in both E5-expressing cell lines compared to empty retrovirus-infected cells, CD1d mRNA levels were not affected. Confocal microscopy demonstrated that residual CD1d was not trafficked to the E5-expressing cell surface but colocalized with E5 near the endoplasmic reticulum (ER). In the ER, E5 interacted with calnexin, an ER chaperone known to mediate folding of CD1d. CD1d protein levels were rescued by the proteasome inhibitor, MG132, indicating a role for proteasome-mediated degradation in HPV-associated CD1d downregulation. Taken together, our data suggest that E5 targets CD1d to the cytosolic proteolytic pathway by inhibiting calnexin-related CD1d trafficking. Finally, CD1d-mediated production of interleukin-12 from the C33A/CD1d cells was abrogated in both E5-expressing cell lines. Decreased CD1d expression in the presence of HPV E5 may help HPV-infected cells evade protective immunological surveillance

Altered Expression of Diabetes-Related Genes in Alzheimer's Disease Brains: The Hisayama Study

Diabetes mellitus (DM) is considered to be a risk factor for dementia including Alzheimer's disease (AD). However, the molecular mechanism underlying this risk is not well understood. We examined gene expression profiles in postmortem human brains donated for the Hisayama study. Three-way analysis of variance of microarray data from frontal cortex, temporal cortex, and hippocampus was performed with the presence/absence of AD and vascular dementia, and sex, as factors. Comparative analyses of expression changes in the brains of AD patients and a mouse model of AD were also performed. Relevant changes in gene expression identified by microarray analysis were validated by quantitative real-time reverse-transcription polymerase chain reaction and western blotting. The hippocampi of AD brains showed the most significant alteration in gene expression profile. Genes involved in noninsulin-dependent DM and obesity were significantly altered in both AD brains and the AD mouse model, as were genes related to psychiatric disorders and AD. The alterations in the expression profiles of DM-related genes in AD brains were independent of peripheral DM-related abnormalities. These results indicate that altered expression of genes related to DM in AD brains is a result of AD pathology, which may thereby be exacerbated by peripheral insulin resistance or DM

A Placebo-Controlled, Double-Blind Randomized (Phase IIB) Trial of Oral Administration with HPV16 E7-Expressing Lactobacillus, GLBL101c, for the Treatment of Cervical Intraepithelial Neoplasia Grade 2 (CIN2)

Cervical intraepithelial neoplasia (CIN), a precursor lesion to cervical cancer, is caused by high-risk human papillomavirus (HPV); high-grade CIN lesions (CIN2-3) are precancerous and require treatment. No globally approved therapy is available for CIN2-3 treatment. This study is a placebo-controlled randomized clinical trial of GLBL101c treatment for CIN2 in 40 patients with HPV16-positive CIN2 who were 1:1 randomized to receive GLBL101c (1 g/daily) or placebo for 5 days at 1, 2, 4, and 8 weeks. No differences were noted between the GLBL101c and placebo groups for patient background and adverse events. Moreover, no statistically significant difference was noted between the two groups at the primary endpoint, pathological regression after 16 weeks of the first oral dose; however, only in the GLBL101c group, two patients had complete regression (CR; regression to normal within 16 weeks). IFNγ production was significantly correlated with the number of spots identified by the interferon gamma enzyme-linked immunospot (IFNγ-ELISPOT) assay using cervical lymphocytes (CxLs) or peripheral blood mononuclear cells. In the two cases of CR, E7-specific Th1 immune responses were observed at week 16. Therefore, we concluded as a novel Lactobacillus-based vaccine with stronger immunogenicity than GLBL101c should be developed

The oncogene KRAS promotes cancer cell dissemination by stabilizing spheroid formation via the MEK pathway

Abstract Background Peritoneal dissemination is a critical prognostic factor in ovarian cancer. Although stabilized spheroid formation promotes cancer cell peritoneal dissemination in ovarian cancer, the associated oncogenes are unknown. In this study, we assessed the role of the KRAS oncogene in ovarian cancer cell dissemination, focusing on the stability of cells in spheroid condition, as well as the modulation of intracellular signaling following spheroid transformation. Methods We used ID8, a murine ovarian cancer cell line, and ID8-KRAS, an oncogenic KRAS (G12 V)-transduced ID8 cell line in this study. Spheroid-forming (3D) culture and cell proliferation assays were performed to evaluate the growth characteristics of these cells. cDNA microarray analysis was performed to identify genes involved in KRAS-associated signal transduction in floating condition. A MEK inhibitor was used to evaluate the effect on cancer peritoneal dissemination. Results Cell viability and proliferation in monolayer (2D) cultures did not differ between ID8 and ID8-KRAS cells. However, the proportions of viable and proliferating ID8-KRAS cells in 3D culture were approximately 2-fold and 5-fold higher than that of ID8, respectively. Spheroid-formation was increased in ID8-KRAS cells. Analysis of peritoneal floating cells obtained from mice intra-peritoneally injected with cancer cells revealed that the proportion of proliferating cancer cells was approximately 2-fold higher with ID8-KRAS than with ID8 cells. Comprehensive cDNA microarray analysis revealed that pathways related to cell proliferation, and cell cycle checkpoint and regulation were upregulated specifically in ID8-KRAS cells in 3D culture, and that some genes partially regulated by the MEK-ERK pathway were upregulated only in ID8-KRAS cells in 3D culture. Furthermore, a MEK inhibitor, trametinib, suppressed spheroid formation in 3D culture of ID8-KRAS cells, although trametinib did not affect 2D-culture cell proliferation. Finally, we demonstrated that trametinib dramatically improved the prognosis for mice with ID8-KRAS tumors in an in vivo mouse model. Conclusions Our data indicated that KRAS promoted ovarian cancer dissemination by stabilizing spheroid formation and that the MEK pathway is important for stabilized spheroid formation. Disruption of spheroid formation by a MEK inhibitor could be a therapeutic target for cancer peritoneal dissemination

Intracellular signaling entropy can be a biomarker for predicting the development of cervical intraepithelial neoplasia.

While the mortality rates for cervical cancer have been drastically reduced after the introduction of the Pap smear test, it still is one of the leading causes of death in women worldwide. Additionally, studies that appropriately evaluate the risk of developing cervical lesions are needed. Therefore, we investigated whether intracellular signaling entropy, which is measured with microarray data, could be useful for predicting the risks of developing cervical lesions. We used three datasets, GSE63514 (histology), GSE27678 (cytology) and GSE75132 (cytology, a prospective study). From the data in GSE63514, the entropy rate was significantly increased with disease progression (normal < cervical intraepithelial neoplasia, CIN < cancer) (Kruskal-Wallis test, p < 0.0001). From the data in GSE27678, similar results (normal < low-grade squamous intraepithelial lesions, LSILs < high-grade squamous intraepithelial lesions, HSILs ≤ cancer) were obtained (Kruskal-Wallis test, p < 0.001). From the data in GSE75132, the entropy rate tended to be higher in the HPV-persistent groups than the HPV-negative group. The group that was destined to progress to CIN 3 or higher had a tendency to have a higher entropy rate than the HPV16-positive without progression group. In conclusion, signaling entropy was suggested to be different for different lesion statuses and could be a useful biomarker for predicting the development of cervical intraepithelial neoplasia