Redox sensor NPGPx restrains ZAP70 activity and modulates T cell homeostasis.

Emerging evidences implicate the contribution of ROS to T cell activation and signaling. The tyrosine kinase, ζ-chain-associated protein of 70 kDa (ZAP70), is essential for T cell development and activation. However, it remains elusive whether a direct redox regulation affects ZAP70 activity upon TCR stimulation. Here, we show that deficiency of non-selenocysteine containing phospholipid hydroperoxide glutathione peroxidase (NPGPx), a redox sensor, results in T cell hyperproliferation and elevated cytokine productions. T cell-specific NPGPx-knockout mice reveal enhanced T-dependent humoral responses and are susceptible to experimental autoimmune encephalomyelitis (EAE). Through proteomic approaches, ZAP70 is identified as the key interacting protein of NPGPx through disulfide bonding. NPGPx is activated by ROS generated from TCR stimulation, and modulates ZAP70 activity through redox switching to reduce ZAP70 recruitment to TCR/CD3 complex in membrane lipid raft, therefore subduing TCR responses. These results reveal a delicate redox mechanism that NPGPx serves as a modulator to curb ZAP70 functions in maintaining T cell homeostasis.

Glutathione peroxidase 8 negatively regulates caspase-4/11 to protect against colitis.

Human caspase-4 and its mouse homolog caspase-11 are receptors for cytoplasmic lipopolysaccharide. Activation of the caspase-4/11-dependent NLRP3 inflammasome is required for innate defense and endotoxic shock, but how caspase-4/11 is modulated remains unclear. Here, we show that mice lacking the oxidative stress sensor glutathione peroxidase 8 (GPx8) are more susceptible to colitis and endotoxic shock, and exhibit reduced richness and diversity of the gut microbiome. C57BL/6 mice that underwent adoptive cell transfer of GPx8-deficient macrophages displayed a similar phenotype of enhanced colitis, indicating a critical role of GPx8 in macrophages. GPx8 binds covalently to caspase-4/11 via disulfide bonding between cysteine 79 of GPx8 and cysteine 118 of caspase-4 and thus restrains caspase-4/11 activation, while GPx8 deficiency leads to caspase-4/11-induced inflammation during colitis and septic shock. Inhibition of caspase-4/11 activation with small molecules reduces the severity of colitis in GPx8-deficient mice. Notably, colonic tissues from patients with ulcerative colitis display low levels of Gpx8 and high caspase-4 expression. In conclusion, these results suggest that GPx8 protects against colitis by negatively regulating caspase-4/11 activity.

Cellular 5-hydroxylmethylcytosine content determines tumorigenic potential and prognosis of pancreatic ductal adenocarcinoma.

We stratified pancreatic ductal adenocarcinoma (PDAC) based on the tumorigenic properties of cancer cells, and aimed to identify clinically useful immunohistochemical (IHC) markers with mechanistic insights. The tumorigenic properties of PDACs were determined using patient-derived xenograft in NOD/SCID/IL2Rγnull mice. The success of tumor engraftment was significantly correlated to poor survival, and its predictive values were superior to clinicopathological parameters. To search IHC-based biomarkers as surrogate for high tumorigenicity with prognostic values, 11 candidates of potentially clinical useful prognostic markers were selected. Among them, 5hmC content of the cancer cells was validated. Elevated 5hmC content positively correlated with in vivo tumorigenicity and poor prognosis in both primary and validation cohorts. Enrichment of cancer-associated 5hmC in CDX2 and FOXA1 lineage-specific transcriptional factor genes further pointed out the potential role of 5hmC in modulating cellular differentiation to enhance tumor malignancy during PDAC progression. Tumor-associated 5hmC content defined a subpopulation of PDAC with high lineage plasticity and tumorigenic potential, and was a prognostic IHC marker that provided a clinical basis for future management of PDAC.

TGF-ß1 secreted by Tregs in lymph nodes promotes breast cancer malignancy via up-regulation of IL-17RB.

Lymph node (LN) metastasis is commonly associated with systemic distant organ metastasis in human breast cancer and is an important prognostic predictor for survival of breast cancer patients. However, whether tumor-draining LNs (TDLNs) play a significant role in modulating the malignancy of cancer cells for distant metastasis remains controversial. Using a syngeneic mouse mammary tumor model, we found that breast tumor cells derived from TDLN have higher malignancy and removal of TDLNs significantly reduced distant metastasis. Up-regulation of oncogenic Il-17rb in cancer cells derived from TDLNs contributes to their malignancy. TGF-ß1 secreted from regulatory T cells (Tregs) in the TDLNs mediated the up-regulation of Il-17rb through downstream Smad2/3/4 signaling. These phenotypes can be abolished by TGF-ß1 neutralization or depletion of Tregs. Consistently, clinical data showed that the up-regulation of IL-17RB in cancer cells from LN metastases correlated with the increased prevalence of Tregs as well as the aggressive growth of tumors in mouse xenograft assay. Together, these results indicate that Tregs in TDLNs play an important role in modulating the malignancy of breast cancer cells for distant metastasis. Blocking IL-17RB expression could therefore be a potential approach to curb the process.

A High Circulating Tumor Cell Count in Portal Vein Predicts Liver Metastasis From Periampullary or Pancreatic Cancer: A High Portal Venous CTC Count Predicts Liver Metastases.

Circulating tumor cells (CTCs) released from a periampullary or pancreatic cancer can be more frequently detected in the portal than the systemic circulation and potentially can be used to identify patients with liver micrometastases. Aims of this study is to determine if CTCs count in portal venous blood of patients with nonmetastatic periampullary or pancreatic adenocarcinoma can be used as a predictor for subsequent liver metastases. CTCs were quantified in portal and peripheral venous blood samples collected simultaneously during pancreaticoduodenectomy in patients with presumed periampullary or pancreatic adenocarcinoma without image-discernible metastasis. Postoperatively patients were monitored for liver metastasis by abdominal magnetic resonance imaging or computed tomography every 3 months for 1 year. Sixty patients with a pathological diagnosis of periampullary or pancreatic adenocarcinoma were included in the study. Multivariate analysis indicated that portal CTC count was a significant predictor for liver metastases within 6 months after surgery. Eleven of 13 patients with a high portal CTCs count (defined as >112 CMx Platform estimated CTCs in 2 mL blood) developed liver metastases within 6 months after surgery. In contrast, only 6 of 47 patients with a low portal CTC count developed liver metastases (P < 0.0001). A value of 112 CMx Platform estimated CTCs had 64.7% sensitivity and 95.4% specificity to predict liver metastases within 6 months after surgery. We concluded that a high CTC count in portal venous blood collected during pancreaticoduodenectomy in patients with periampullary or pancreatic adenocarcinoma without metastases detected by currently available imaging tools is a significant predictor for liver metastases within 6 months after surgery.

RANKL/RANK control Brca1 mutation- .

Breast cancer is the most common female cancer, affecting approximately one in eight women during their life-time. Besides environmental triggers and hormones, inherited mutations in the breast cancer 1 (BRCA1) or BRCA2 genes markedly increase the risk for the development of breast cancer. Here, using two different mouse models, we show that genetic inactivation of the key osteoclast differentiation factor RANK in the mammary epithelium markedly delayed onset, reduced incidence, and attenuated progression of Brca1;p53 mutation-driven mammary cancer. Long-term pharmacological inhibition of the RANK ligand RANKL in mice abolished the occurrence of Brca1 mutation-driven pre-neoplastic lesions. Mechanistically, genetic inactivation of Rank or RANKL/RANK blockade impaired proliferation and expansion of both murine Brca1;p53 mutant mammary stem cells and mammary progenitors from human BRCA1 mutation carriers. In addition, genome variations within the RANK locus were significantly associated with risk of developing breast cancer in women with BRCA1 mutations. Thus, RANKL/RANK control progenitor cell expansion and tumorigenesis in inherited breast cancer. These results present a viable strategy for the possible prevention of breast cancer in BRCA1 mutant patients.

Developmental phenotype of a membrane only estrogen receptor alpha (MOER) mouse.

Estrogen receptors (ERs) alpha and beta exist as nuclear, cytoplasmic, and membrane cellular pools in a wide variety of organs. The relative contributions of each ERalpha pool to in vivo phenotypes resulting from estrogen signaling have not been determined. To address this, we generated a transgenic mouse expressing only a functional E domain of ERalpha at the plasma membrane (MOER). Cells isolated from many organs showed membrane only localized E domain of ERalpha and no other receptor pools. Liver cells from MOER and wild type mice responded to 17-beta-estradiol (E2) with comparable activation of ERK and phosphatidylinositol 3-kinase, not seen in cells from ERalphaKO mice. Mating the MOER female mice with proven male wild type breeders produced no pregnancies because the uterus and vagina of the MOER female mice were extremely atrophic. Ovaries of MOER and homozygous Strasbourg ERalphaKO mice showed multiple hemorrhagic cysts and no corpus luteum, and the mammary gland development in both MOER and ERalphaKO mice was rudimentary. Despite elevated serum E2 levels, serum LH was not suppressed, and prolactin levels were low in MOER mice. MOER and Strasbourg female mice showed plentiful abdominal visceral and other depots of fat and increased body weight compared to wild type mice despite comparable food consumption. These results provide strong evidence that the normal development and adult functions of important organs in female mice requires nuclear ERalpha and is not rescued by membrane ERalpha domain expression alone.