Notch signaling is activated by TLR stimulation and regulates macrophage functions

Notch signaling is a well-conserved pathway involved in cell fate decisions, proliferation and apoptosis. We report on the involvement of Notch signaling in regulating gene expression in activated macrophages. Toll-like receptors (TLR) agonists such as bacterial lipopeptide, polyI:C, lipopolysaccharide and unmethylated CpG DNA all induced up-regulation of Notch1 in primary and macrophage-like cell lines. Notch1 up-regulation was dependent on the MyD88 pathway when stimulated through TLR2, but not TLR4. Activated Notch1 and expression of the Notch target genes, Hes1 and Deltex, were detected in activated macrophages, suggesting that Notch signaling was activated upon stimulation. Inhibiting processing of Notch receptor by gamma-secretase using a gamma-secretase inhibitor (GSI), the expression of Notch1 was down-regulated to basal levels. This treatment significantly modulated expression of TNF-alpha, IL-6, and IL-10. In addition, the amount of nitric oxide produced was significantly lower and the expression of MHC class II was up-regulated in GSI-treated cells. Treatment with GSI or silencing Notch1 resulted in decreased translocation of NF-kappaBp50 into nucleus upon stimulation. Taken together, stimulation of macrophages through the TLR signaling cascade triggered activation of Notch signaling, which in turn regulated gene expression patterns involved in pro-inflammatory responses, through activation of NF-kappaB.     

Impact of Obesity on Outcomes of Operable Breast Cancer: A Retrospective Cohort Study

Objective: Obesity is increasing worldwide. Previous studies of the impact of obesity on breast cancer outcomes have reported conflicting results. We investigated the association of obesity and breast cancer survival in Thai patients. Methods: Medical records of operable breast cancer patients diagnosed and treated at Siriraj Hospital between January 2004 and December 2011 were reviewed. Demographic data, tumor characteristics, stage, treatment and adverse event were described. Obesity was defined as body mass index (BMI) ≥ 25 kg/m2 using Asian’s cutoff value.  Survivals in both obese and non-obese patient groups were analyzed. Results: A total of 400 patients were included, 200 in each group. Obese patients were older and associated with more comorbidity. Obesity was associated with larger tumor size (p = 0.011), greater numbers of lymph node involvement (p = 0.003) and more advanced stage (p = 0.01). Obese patients were more likely to receive less adjuvant chemotherapy and hormonal treatment. There was no statistically significant difference in disease-free survival (DFS) (Hazard ratio [HR] 0.72, 95% confidence interval [CI] 0.46 to 1.13) and overall survival (OS) (HR 0.77, 95% CI 0.43 to 1.39) between obese and non-obese patients. Interestingly, obesity was associated with fewer complications from chemotherapy than non-obese patients (p = 0.047). Conclusion: Obesity had no adverse prognostic impact association on both DFS and OS in Thai patients with operable breast cancer, although obese patients more often presented with larger tumor and higher numbers of lymph node involvement.     

Hippocampal neurogenesis in adult Old World primates

The production of new hippocampal neurons in adulthood has been well documented in rodents. Recent studies have extended these findings to other mammalian species, such as tree shrews and marmoset monkeys. However, hippocampal neurogenesis has not been demonstrated in adult Old World primates. To investigate this possibility, we injected 11 adult Old World monkeys of different ages (5-23 years) with the thymidine analog bromodeoxyuridine and examined the fate of the labeled cells at different survival times by using neuronal and glial markers. In the young-adult and middle-aged monkeys, we found a substantial number of cells that incorporated bromodeoxyuridine and exhibited morphological and biochemical characteristics of immature and mature neurons. New cells located in the dentate gyrus expressed a marker of immature granule neurons, Turned On After Division 64 kDa protein, as well as markers of mature granule neurons including neuron specific enolase, neuronal nuclei, and the calcium-binding protein calbindin. Fewer new cells expressed the astroglial marker glial fibrillary acidic protein. Evidence of neurogenesis was observed in the oldest monkeys (23 years) as well, but it appeared to be less robust. These results indicate that the adult brains of Old World monkeys produce new hippocampal neurons. Adult macaque monkeys may provide a useful primate model for studying the functional significance of adult neurogenesis.     

Regulation of hippocampal neurogenesis in adulthood.

A substantial number of new granule neurons are produced in the dentate gyrus in adulthood in a variety of mammalian species, including humans. Numerous studies have demonstrated that the production and survival of new hippocampal neurons can be enhanced or diminished by hormones and experience. Steroid hormones of the ovaries and adrenal glands have been shown to modulate the production of immature neurons by affecting the proliferation of granule cell precursors. Aversive experiences have been demonstrated to decrease the production of immature granule cells, whereas enriching experiences, including learning, have been shown to enhance the survival of new hippocampal cells. These studies indicate that adult-generated neurons represent a unique form of structural plasticity that can be regulated by the environment, and furthermore suggest that new neurons play an important role in hippocampal function.     

Enhancement of immune response to a DNA vaccine against Mycobacterium tuberculosis Ag85B by incorporation of an autophagy inducing system

DNA vaccines are a promising new generation of vaccines that can elicit an immune response using DNA encoding the antigen of interest. The efficacy of these vaccines, however, still needs to be improved. In this study, we investigated the effect of autophagy on increasing the efficacy of a candidate DNA vaccine against Mycobacterium tuberculosis (MTB), a causative agent of tuberculosis. Low molecular weight chitosan was used to encapsulate plasmid DNA containing a gene encoding MTB Antigen 85B (Ag85B), a secreted fibronectin-binding protein. To induce autophagy upon DNA vaccination, the kinase defective mTOR (mTOR-KD) was transfected into cells, and autophagy was detected based on the presence of LC3II. To investigate whether autophagy enhances an immune response upon DNA vaccination, we coencapsualted the Ag85B-containing plasmid with a plasmid encoding mTOR-KD. Plasmids encapsulated by chitosan particles were used for primary subcutaneous immunization and for intranasal boost in mice. After the boost vaccination, sera from the mice were measured for humoral immune response. The DNA vaccine with the autophagy-inducing construct elicited significantly higher Ag85B-specific antibody levels than the control group treated with the Ag85B plasmid alone or with the Ag85B plasmid plus the wild type mTOR construct. Upon in vitro stimulation of splenocytes from mice immunized with recombinant Ag85B, the highest levels of secreted IFN-γ and IL-2 were detected in mice immunized with the autophagy-inducing plasmid, while no differences in IL-4 levels were detected between the groups, suggesting that the DNA vaccine regimen with autophagy induction induced primarily a Th1 immune response. Furthermore, the enhanced proliferation of CD4+ T cells from mice receiving the autophagy-inducing vaccine was observed in vitro. Based on the evidence presented, we conclude that incorporating an autophagy-inducing element into a DNA vaccine may help to improve immune response.