Relationship between menopausal hormone therapy and incidence of fractures in postmenopausal women.

OBJECTIVE: Long-term protective effects of menopausal hormone therapy (MHT) at fractures with different doses and components are controversial. We analyzed the effect of MHT on the incidence of spine and femur fractures according to MHT type, age at commencement, duration and dose of hormones in Korean women. METHOD: This retrospective study evaluated propensity score-matched patients with MHT from the Korean National Health Insurance Service database. Among women aged ≥50 years with menopause between 2004 and 2007, spine and femur fracture incidence until 2017 was analyzed in 36,446 women who had received MHT for >1 year. Estrogen-progesterone therapy (EPT), estrogen-only therapy (ET) or tibolone therapy was conducted. RESULTS: EPT significantly lowered the incidence of spine and femur fractures with a conventional dose, but not with a low dose. Tibolone significantly decreased the incidence of spine fractures in women aged 50-59 years when used for >5 years, and the incidence of femur fractures in women older than 60 years when used for >3 years. ET significantly lowered the risk of femur fractures when estradiol was used for >5 years. CONCLUSION: In menopausal women, all MHT including conventional-dose EPT, ET and tibolone tended to lower the incidence of fractures. The effects, however, varied with the type of fracture and type of MHT.

Antibody-mediated blockade for galectin-3 binding protein in tumor secretome abrogates PDAC metastasis.

The major challenges in pancreatic ductal adenocarcinoma (PDAC) management are local or distant metastasis and limited targeted therapeutics to prevent it. To identify a druggable target in tumor secretome and to explore its therapeutic intervention, we performed a liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic analysis of tumors obtained from a patient-derived xenograft model of PDAC. Galectin-3 binding protein (Gal-3BP) is identified as a highly secreted protein, and its overexpression is further validated in multiple PDAC tumors and primary cells. Knockdown and exogenous treatment of Gal-3BP showed that it is required for PDAC cell proliferation, migration, and invasion. Mechanistically, we revealed that Gal-3BP enhances galectin-3-mediated epidermal growth factor receptor signaling, leading to increased cMyc and epithelial-mesenchymal transition. To explore the clinical impact of these findings, two antibody clones were developed, and they profoundly abrogated the metastasis of PDAC cells in vivo. Altogether, our data demonstrate that Gal-3BP is an important therapeutic target in PDAC, and we propose its blockade by antibody as a therapeutic option for suppressing PDAC metastasis.

Identification of Single Nucleotide Polymorphisms as Biomarkers for Recurrent Pregnancy Loss in Korean Women.

BACKGROUND: Single nucleotide polymorphisms (SNPs) are reportedly associated with repeated abortion. Thus, genetic analysis based on race is the key to developing accurate diagnostic tests. This study analyzed the genetic polymorphisms of recurrent pregnancy loss (RPL) patients among Korean women compared to the controls. METHODS: In 53 women of RPL group and 50 controls, the genetic analysis was performed. The genotype distribution and allele frequency were analyzed statistically for the difference between the two groups. The association between each SNP marker and RPL risk was analyzed. RESULTS: The genotypes of LEPR, endothelial nitric oxide synthase (eNOS), KDR, miR-27a, miR-449b, and tumor necrosis factor-alpha (TNF-α) were analyzed using odds ratio (OR) with 95% confidence intervals (CIs). Only the AG genotype of miR-449b (A>G) polymorphism showed significant association with the risk of RPL when compared to the AA genotype (OR, 2.39). The combination of GG/AG+GG/CA+AA genotypes for eNOS/miR-449b/TNF-α was associated with 7.36-fold higher risk of RPL (OR, 7.36). The GG/AG+GG combination for eNOS/miR-449b showed 2.43-fold higher risk for RPL (OR, 2.43). The combination of AG+GG/CA+AA genotypes for miR-449b/TNF-α showed a significant association with the risk of RPL (OR, 7.60). From the haplotype-based analysis, the G-G-A haplotype of eNOS/miR-449b/TNF-α and the G-A haplotype of miR-449b/TNF-α were associated with increased risk of RPL (OR, 19.31; OR, 22.08, respectively). CONCLUSION: There is a significant association between the risk of RPL and miR-449b/TNF-α combination, and therefore, genetic analysis for specific combined genotypes can be an important screening method for RPL in Korean women.

Metformin use in cancer survivors with diabetes reduces all-cause mortality, based on the Korean National Health Insurance Service between 2002 and 2015.

ABSTRACT: Malignant neoplasms are the leading cause of death in Korea. We aimed to examine if metformin use in cancer survivors reduces all-cause mortality. This study was retrospectively designed based on data from the Korean National Health Insurance Service-National Health Screening Cohort (HEALS) between 2002 and 2015. The Kaplan-Meier estimator and log-rank test was performed to estimate the survival function according to metformin usage (3721 metformin non-users with diabetes, 5580 metformin users with diabetes, and 24,483 non-diabetic individuals). Adjusted hazard ratios (HRs) and 95% confidence intervals (CIs) for all-cause mortality were calculated using Cox proportional hazards regression models.The median follow-up duration was 4.2 years. The HRs (95% CIs) for all-cause mortality of metformin users and the non-diabetic group were 0.762 (0.683-0.850) and 1.055 (0.966-1.152) in men and 0.805 (0.649-0.999), and 1.049 (0.873-1.260) in women, respectively, compared with metformin non-users among diabetic cancer survivors, in a fully adjusted model. After stratifying metformin users into pre- and post-diagnosis of cancers, adjusted HRs (95% CIs) of pre- and post-diagnosis metformin users for all-cause mortality were 0.948 (0.839-1.071) and 0.530 (0.452-0.621) in men and 1.163 (0.921-1.469) and 0.439 (0.323-0.596) in women, respectively.Metformin use in cancer survivors with diabetes reduced overall mortality rates. In particular, metformin use after cancer diagnosis, not before cancer diagnosis, was inversely associated with overall mortality.Active treatment with metformin for diabetic cancer survivors after cancer diagnosis can improve their survival rates.

Metformin usage and the risk of colorectal cancer: a national cohort study.

PURPOSE: This study aimed to investigate the association between metformin usage and the risk of colorectal cancer (CRC) using data from the Korean National Health Insurance Service-National Health Screening Cohort database. METHODS: Data from the NHIS-HEALS cohort between 2002 and 2015 were longitudinally analyzed. Subjects were divided into three groups: metformin non-users with diabetes mellitus (DM), metformin users with DM, and no DM group. CRC was defined using the ICD-10 code (C18.0-C20.0) at the time of admission. Cox proportional hazard regression models were adopted after stepwise adjustment for confounders to investigate the association between metformin usage and colorectal cancer risk. RESULTS: During the follow-up period, of the total 323,430 participants, 2341 (1.33%) of the 175,495 males and 1204 (0.81%) of the 147,935 females were newly diagnosed with CRC. The estimated cumulative incidence of CRC was significantly different among the three groups based on Kaplan-Meier's survival curve (p values < 0.05 in both sexes). Compared with metformin non-users, hazard ratios (95% CIs) of metformin users and the no DM group were 0.66 (0.51-0.85) and 0.72 (0.61-0.85) in males and 0.59 (0.37-0.92) and 0.93 (0.66-1.29) in females, respectively, after being fully adjusted. CONCLUSIONS: Metformin users with diabetes appear to have a significantly lower risk of CRC compared with metformin non-users.

Metformin use reduced the overall risk of cancer in diabetic patients: A study based on the Korean NHIS-HEALS cohort.

BACKGROUND AND AIM: Several studies have reported the preventive effect of metformin on cancer development. This study aimed to investigate the relationship between use of metformin and risk of cancer in Koreans. METHODS AND RESULTS: This study was designed retrospectively using the National Health Insurance Service-National Health Screening Cohort conducted between 2002 and 2015. 40 to 69-year-old subjects who received a health screening examination from 2002 to 2003 were enrolled. Hazard ratios (HRs) and 95% confidence intervals (CIs) for cancer were estimated in a multivariate Cox proportional regression analysis. A total of 323,430 subjects was enrolled (301,905 individuals without diabetes [No DM], 8643 diabetic patients with metformin treatment [metformin users], and 12,882 diabetic patients without metformin treatment [metformin non-users]). The median follow-up period was 12.7 years. Cumulative incidence of overall cancer was 7.9% (7.7, 10.3, and 11.1% in No DM, metformin users and non-users, respectively). Compared to metformin non-users, the fully adjusted HRs (95% CIs) of metformin users and No DM for overall cancer incidence were 0.73 (0.66-0.81) and 0.75 (0.64-0.88), respectively, in men and 0.83 (0.78-0.89) and 0.81 (0.72-0.92) in women. CONCLUSIONS: Diabetic patients receiving metformin treatment, and individuals without diabetes were at lower risk for cancer incidence than diabetic patients without metformin treatment.

Diabetes, Metformin, and Lung Cancer: Retrospective Study of the Korean NHIS-HEALS Database.

BACKGROUND: Metformin is the first option in managing type 2 diabetes mellitus (DM) and has pleotropic effects. We studied the incidence of lung cancer in patients who received metformin therapy. PATIENTS AND METHODS: This study was retrospectively designed and based on the Korean National Health Insurance Service-National Health Screening Cohort to determine whether metformin reduces lung cancer risk in the diabetic population. At baseline, all participants were 40 to 69 years old and were categorized into 3 groups: metformin nonrecipients with DM, metformin recipients with DM, and the nondiabetic group. RESULTS: A total of 336,168 individuals were included in the final analysis (314,291 nondiabetic individuals, 8806 metformin recipients, and 13,071 metformin nonrecipients). The study median follow-up period was 12.86 years. The estimated cumulative lung cancer incidence of metformin nonrecipients, metformin recipients, and the nondiabetic group was 1.80%, 1.97%, and 1.24% in men and 1.87%, 0.61%, and 0.41% in women, respectively (P < .05). Compared to metformin nonrecipients, the hazard ratios (95% confidence intervals) for lung cancer incidence of metformin recipients and the nondiabetic group were 1.287 (0.979-1.691) and 0.835 (0.684-1.019) in men and 0.664 (0.374-1.177) and 0.553 (0.359-0.890) in women, respectively. The hazard ratios (95% confidence intervals) were statistically significant in male ever smokers (0.784 [0.627-0.979]) and female nonsmokers (0.498 [0.320-0.774]) after stratification according to smoking status. CONCLUSION: Metformin therapy did not reduce lung cancer incidence in the diabetic population. However, individuals without DM were at a lower risk of lung cancer, especially in male ever smokers and female nonsmokers.

Metformin use reduced the risk of stomach cancer in diabetic patients in Korea: an analysis of Korean NHIS-HEALS database.

BACKGROUND: Diabetes mellitus (DM) increases atherosclerotic cardiovascular complications and cancer risks. Stomach cancer is the most common cancer in Korea. Although the survival rate of stomach cancer has improved, the disease burden is still high. METHODS: This retrospective study investigated the association between metformin use and stomach cancer incidence in a Korean population using the National Health Insurance Service-National Health Screening Cohort database. Participants aged 40-80 years old at the baseline period (2002-2003) were enrolled. The study population was categorized into three groups of metformin non-users with DM, metformin users with DM, and individuals without DM (No DM group). RESULTS: A total of 347,895 participants (14,922 metformin non-users, 9891 metformin users, and 323,082 individuals without DM) were included in the final analysis. The median follow-up duration was 12.70 years. The estimated cumulative incidence of stomach cancer was highest in metformin non-users and lowest in the No DM group (men vs. women: 3.75 vs. 1.97% in metformin non-users, 2.91 vs. 1.53% in metformin users, and 2.54 vs. 0.95% in the No DM group). Compared with metformin non-users, the hazard ratios (95% confidence intervals) for stomach cancer incidence of metformin users and the No DM group were 0.710 (0.579-0.870) and 0.879 (0.767-1.006) in men and 0.700 (0.499-0.981) and 0.701 (0.544-0.903) in women, respectively, after full adjustment. CONCLUSIONS: Metformin users with DM in the Korean population were at lower risk of stomach cancer incidence after controlling for potential confounding factors.

Statin use increased new-onset diabetes in hypercholesterolemic individuals: Data from the Korean National Health Insurance Service-National Health Screening Cohort database (NHIS-HEALS).

AIM: We investigated the association between statin use and new-onset diabetes (NODM) in Korean adults with hypercholesterolemia. METHODS: This study performed based on data from the National Health Insurance Service-National Health Screening Cohort for the years from 2002 to 2015. Statin users classified as high- or low- users according to medication possession ratio. Statin non-users consisted of hypercholesterolemic participants who never used statin over the entire follow-up period. 21,469 participants (10,880 statin users, 10,589 statin non-users) with a median follow-up period of 12.5 years were included. We estimated the NODM risk based on the survival analyses. In particular, to adjust for confounding effects, we considered Cox proportional hazards regression models over three stages. RESULTS: Compared to non-users, statin users had a significantly higher risk for NODM. The fully adjusted hazard ratios (aHRs) (95% confidential intervals [95% CIs]) of statin users for NODM were 1.43 (1.31-1.57) in men, and 1.86 (1.66-2.10) in women, respectively after adjusted confounding factors including age and lifestyle factors. Compared to high-users, aHRs (95% CIs) of low-users for NODM were 1.16 (1.03-1.30) and 1.28 (1.16-1.43) in men and women, respectively. CONCLUSIONS: In hypercholesterolemic patients, statin users have a higher risk of NODM than non-users.

Association between statin use and all-cause mortality in cancer survivors, based on the Korean health insurance service between 2002 and 2015.

BACKGROUND AND AIMS: Cancer is the number one cause of death in Korea. This study aimed to investigate if statin use in cancer survivors was inversely associated with all-cause mortality. METHODS AND RESULTS: Data from the 2002 to 2015 National Health Insurance Service-National Health Screening Cohort (NHIS-HEALS) were used. The Kaplan-Meier estimator was used to estimate the survival function according to statin usage. Cox proportional hazards regression models were adopted after stepwise adjustment for potential confounders to calculate hazard ratios (HRs) and 95% confidence intervals (CIs) for all-cause mortality. The median follow-up duration was 10.0 years. Statin users had a higher percentage of diabetes and hypertension in both sexes. Survival rates of statin users were higher than non-users (p-values <0.001 in men and 0.021 in women). Compared to non-users, the HRs (95% CIs) of statin users for all-cause mortality were 0.327 (0.194-0.553) in men and 0.287 (0.148-0.560) in women after adjustment for potential confounding factors. CONCLUSIONS: Statin users in cancer survivors had higher survival rate than non-users in both sexes.

A pharmacogenomic analysis using L1000CDS2 identifies BX-795 as a potential anticancer drug for primary pancreatic ductal adenocarcinoma cells.

Pancreatic cancer is one of the leading causes of cancer death, mainly due to the absence of early diagnostic tool and effective therapeutic agents. To identify an effective therapeutic agent for pancreatic ductal adenocarcinoma cells (PDAC), we used 10 Gene Expression Omnibus (GEO) data sets and L1000CDS2 pharmacogenetic search tool and obtained chemical "perturvants" that were predicted to reverse the abnormal gene expression changes in PDAC. Among 20 initial candidates, we measured IC50 for six compounds and identified BX-795, PDK1/TBK1 inhibitor, as a therapeutic candidate. We found that BX-795 inhibits primary PDAC cell proliferation more effectively than normal cells. Following molecular analysis revealed that BX-795 down-regulates mTOR-GSK3ß pathway and trigger apoptosis. Moreover, we found that BX-795 suppresses primary PDAC cell migration via downregulation of Snail and Slug. Finally, efficacy test in patient-derived xenograft model of PDAC showed BX-795 can inhibit in vivo tumor growth as efficient as gemcitabine and a combination with trametinib further suppresses tumor growth. Collectively, these results demonstrate the BX-795 as an effective therapeutic candidate for PDAC treatment.

Paraventricular Thalamus Controls Behavior during Motivational Conflict.

Decision-making often involves motivational conflict because of the competing demands of approach and avoidance for a common resource: behavior. This conflict must be resolved as a necessary precursor for adaptive behavior. Here we show a role for the paraventricular thalamus (PVT) in behavioral control during motivational conflict. We used Pavlovian counterconditioning in male rats to establish a conditioned stimulus (CS) as a signal for reward (or danger) and then transformed the same CS into a signal for danger (or reward). After such training, the CS controls conflicting appetitive and aversive behaviors. To assess PVT involvement in conflict, we injected an adeno-associated virus (AAV) expressing the genetically encoded Ca2+ indicator GCaMP and used fiber photometry to record population PVT Ca2+ signals. We show distinct profiles of responsivity across the anterior-posterior axis of PVT during conflict, including an ordinal relationship between posterior PVT CS responses and behavior strength. To study the causal role of PVT in behavioral control during conflict, we injected AAV expressing the inhibitory hM4Di DREADD and determined the effects of chemogenetic PVT inhibition on behavior. We show that chemogenetic inhibition across the anterior-posterior axis of the PVT, but not anterior or posterior PVT alone, disrupts arbitration between appetitive and aversive behaviors when they are in conflict but has no effect when these behaviors are assessed in isolation. Together, our findings identify PVT as central to behavioral control during motivational conflict.SIGNIFICANCE STATEMENT Animals, including humans, approach attractive stimuli and avoid aversive ones. However, they frequently face conflict when the demands of approach and avoidance are incompatible. Resolution of this conflict is fundamental to adaptive behavior. Here we show a role for the paraventricular thalamus, a nucleus of the dorsal midline thalamus, in the arbitration of appetitive and aversive behavior during motivational conflict.

A drug-repositioning screen for primary pancreatic ductal adenocarcinoma cells identifies 6-thioguanine as an effective therapeutic agent for TPMT-low cancer cells.

Pancreatic cancer is one of the most difficult cancers to cure due to the lack of early diagnostic tools and effective therapeutic agents. In this study, we aimed to isolate new bioactive compounds that effectively kill pancreatic ductal adenocarcinoma (PDAC) cells, but not untransformed, human pancreatic ductal epithelial (HPDE) cells. To this end, we established four primary PDAC cell lines and screened 4141 compounds from four bioactive-compound libraries. Initial screening yielded 113 primary hit compounds that caused over a 50% viability reduction in all tested PDAC cells. Subsequent triplicate, dose-dependent analysis revealed three compounds with a tumor cell-specific cytotoxic effect. We found that these three compounds fall into a single category of thiopurine biogenesis. Among them, 6-thioguanine (6-TG) showed an IC50 of 0.39-1.13 µm toward PDAC cells but had no effect on HPDE cells. We propose that this cancer selectivity is due to differences in thiopurine methyltransferase (TPMT) expression between normal and cancer cells. This enzyme is responsible for methylation of thiopurine, which reduces its cytotoxicity. We found that TPMT levels were lower in all four PDAC cell lines than in HPDE or Panc1 cells, and that knockdown of TPMT in HPDE or Panc1 cells sensitized them to 6-TG. Lastly, we used a patient-derived xenograft model to confirm that 6-TG has a significant antitumor effect in combination with gemcitabine. Overall, our study presents 6-TG as a strong candidate for use as a therapeutic agent against PDAC with low levels of TPMT.

Induction of apoptosis by sarijang, a bamboo salt sauce, in U937 human leukemia cells through the activation of caspases.

Sarijang is a bamboo salt soy sauce, containing extracts of Rhynchosia nulubilis, sulfur-fed duck, dried bark of Ulmus davidiana and Allium sativum, which has been demonstrated to exert anti-inflammatory and antitumor activity. However, the cellular and molecular mechanisms of action of sarijang have not yet been elucidated. In the present study, we investigated the pro-apoptotic effects of sarijang in an in vitro U937 human leukemia cell model. Treatment with sarijang resulted in a concentration-dependent growth inhibition of the cells, coupled with the characteristic morphological features of apoptosis. The induction of the apoptotic cell death of the U937 cells by sarijang exhibited a correlation with the upregulation of death receptor 4 (DR4), the downregulation of members of the inhibitor of apoptosis protein (IAP) family, including survivin and cellular IAP (cIAP)-1, and the cleavage of Bid. Apoptosis-inducing concentrations of sarijang also induced the activation of caspases (caspase-3, -8 and -9), accompanied by proteolytic degradation of poly(ADP-ribose)-polymerase, ß-catenin and phospholipase C-γ1. However, the apoptosis induced by sarijang was significantly inhibited by z-VED-fmk, a pan-caspase inhibitor, which demonstrated the importance of caspases in the process. These results suggested that sarijang may be a potential chemotherapeutic agent for use in the control of U937 human leukemia cells. Further studies are required to identify the active compounds in sarijang.

Anti-inflammatory effects of egg white combined with chalcanthite in lipopolysaccharide-stimulated BV2 microglia through the inhibition of NF-κB, MAPK and PI3K/Akt signaling pathways.

Egg white-chalcanthite (EWCC) is a mixture of egg white and chalcanthite prepared by roasting chalcanthite (which is a natural mineral mainly composed of CuSO4•5H2O) to the point of dehydration, pulverizing the dehydrated chalcanthite and then mixing the pulverized chalcanthite to react with egg white to trigger a reaction. When egg white-chalcanthite is prepared in this manner, the toxicity of chalcanthite is neutralized by the egg white, so that the toxicity is reduced or removed and the pharmaceutical properties are increased. However, the cellular and molecular mechanisms underlying the pharmacological activity of EWCC remain poorly understood. In this study, we investigated the inhibitory effects of EWCC on the production of lipopolysaccharide (LPS)-induced pro-inflammatory mediators in BV2 microglia. Our data indicated that the EWCC treatment significantly inhibited the excessive production of nitric oxide and prostaglandin E2 in LPS-stimulated BV2 microglia in a concentration-dependent manner without causing cytotoxicity. It also attenuated the expression of inducible nitric oxide synthase, cyclooxygenase-2 and pro-inflammatory cytokines, including interleukin-1ß and tumor necrosis factor-α. Moreover, EWCC exhibited anti-inflammatory properties by the suppression of nuclear factor­κB (NF-κB) activation by blocking IκB-α degradation, downregulation of extracellular signal-regulated kinase, c-Jun N-terminal kinase, p38 mitogen-activated protein kinase (MAPK) and phosphoinositide 3-kinase (PI3K)/Akt pathways. Our results indicate that the inhibitory effects of EWCC on LPS-stimulated inflammatory mediator production in BV2 microglia are associated with the suppression of the NF-κB, MAPK and PI3K/Akt signaling pathways. These findings suggest that EWCC may offer a substantial therapeutic potential for the treatment of neurodegenerative diseases that are accompanied by microglial activation.

Effect of Egg White Combined with Chalcanthite on Lipopolysaccharide induced Inflammatory Cytokine Expression in RAW 264.7 cells.

AIM: Historically, mineral compound herbal medicines have long been used in treatments of immune-related diseases in Korea, China and other Asian countries. In this study, we investigated the anti-inflammatory effect of egg white combined with chalcanthite (IS4) on lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. METHODS: RAW 264.7 cells cultured with LPS and various concentrations of IS4 were analyzed to determine the production of pro-inflammatory cytokines and mediators by using enzyme-linked immune sorbent assays (ELISAs). RESULTS: IS4 concentration inhibited the production of interleukin-1beta (IL-1 ß), interleukin-6 (IL-6), and granulocyte -macrophage colony-stimulating factor (GM-CSF) induced by LPS. IS4 at high concentrations (25 and 50㎍/ml) inhibited, in concentration-dependent manner, the expression of tumor necrosis factor-alpha (TNF- α) stimulated by LPS. CONCLUSION: IS4 has shown an anti-inflammatory effect in RAW 264.7 cells.

Pre-mRNA 3'-end processing complex assembly and function.

The 3'-ends of almost all eukaryotic mRNAs are formed in a two-step process, an endonucleolytic cleavage followed by polyadenylation (the addition of a poly-adenosine or poly(A) tail). These reactions take place in the pre-mRNA 3' processing complex, a macromolecular machinery that consists of more than 20 proteins. A general framework for how the pre-mRNA 3' processing complex assembles and functions has emerged from extensive studies over the past several decades using biochemical, genetic, computational, and structural approaches. In this article, we review what we have learned about this important cellular machine and discuss the remaining questions and future challenges.

Paraventricular thalamus mediates context-induced reinstatement (renewal) of extinguished reward seeking.

Paraventricular thalamus (PvTh) is uniquely placed to contribute to reinstatement of drug and reward seeking. It projects extensively to regions implicated in reinstatement including accumbens shell (AcbSh), prefrontal cortex (PFC) and basolateral amygdala (BLA), and receives afferents from other regions important for reinstatement such as lateral hypothalamus. We used complementary neuroanatomical and functional approaches to study the role of PvTh in context-induced reinstatement (renewal) of extinguished reward-seeking. Rats were trained to respond for a reward in context A, extinguished in context B and tested in context A or B. We applied the neuronal tracer cholera toxin B subunit (CTb) to AcbSh and examined retrograde-labelled neurons, c-Fos immunoreactivity (IR) and dual c-Fos/CTb labelled neurons in PvTh and other AcbSh afferents. In PvTh there was c-Fos IR in CTb-positive neurons associated with renewal showing activation of a PvTh-AcbSh pathway during renewal. In PFC there was little c-Fos IR in CTb-positive or negative neurons associated with renewal. In BLA, two distinct patterns of activation and retrograde labelling were observed. In rostral BLA there was significant c-Fos IR in CTb-negative neurons associated with renewal. In caudal BLA there was significant c-Fos IR in CTb-positive neurons associated with being tested in either the extinction (ABB) or training (ABA) context. We then studied the functional role of PvTh in renewal. Excitotoxic lesions of PvTh prevented renewal. These lesions had no effect on the acquisition of reward seeking. These results show that PvTh mediates context-induced reinstatement and that this renewal is associated with recruitment of a PvTh-AcbSh pathway.

Id-1 gene downregulation by sulindac sulfide and its upregulation during tumor development in gastric cancer.

The molecular mechanisms underlying the chemopreventive effects of NSAIDs are not well understood and remain the subject of debate. One of the mechanistic possibilities involves alterations in gene expression. We examined gene expression profiles in SNU601 gastric cancer cells treated with sulindac sulfide (50 microM) for 24 hr. Microarray analysis showed that 1.3% (105/8170) of genes were induced or repressed more than 3-fold in cells treated with sulindac sulfide. Seven genes were selected and confirmed by reverse transcription-polymerase chain reaction. Inhibitor of differentiation/DNA binding-1 (Id-1) was downregulated in SNU601 cells treated with sulindac sulfide. Id-1 expression level was decreased dose-dependently by sulindac sulfide. In addition, the expression pattern of Id-1 was inversely related to that of nm23. We also examined Id-1 expression in human gastric cancer tissues and compared it with clinicopathologic parameters to study its biologic role in the cancers. Id-1 was frequently and strongly expressed in gastric cancer tissues compared with that in adjacent nonmetaplastic mucosa. Its immunoreactivity scores were positively correlated to Ki67 labeling indices and tumor progression, and is higher in intestinal type than in diffuse type. In summary, a number of genes, both induced and repressed, could be important in mediating sulindac sulfide-induced cell death in gastric cancer cells. Id-1, one of the repressed genes, is upregulated in gastric cancers and has positive role in tumor progression and histogenesis of intestinal-type cancers.

The midbrain periaqueductal gray and fear extinction: opioid receptor subtype and roles of cyclic AMP, protein kinase A, and mitogen-activated protein kinase.

Four experiments studied the opioid receptor subtype and signal transduction mechanisms mediating fear extinction in the ventrolateral quadrant of the midbrain periaqueductal gray (vlPAG). Microinjection of a mu- but not a delta- or kappa-opioid receptor antagonist into the vlPAG retarded extinction. Extinction was also dose-dependently retarded by vlPAG infusions of a cyclic AMP (cAMP) analog but was unaffected by infusions of a protein kinase A activator or a mitogen-activated protein kinase inhibitor across wide dose ranges. The results show that fear extinction occurs via activation of vlPAG mu-opioid receptors and involves reductions in cAMP. These mechanisms are different from the cellular mechanisms for extinction in the amygdala and from the known cellular mechanisms for opioid analgesia in the vlPAG.